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3

 Learning Center

Micro-Volume Sampling—How it Works

176

NanoDrop One User Guide

Thermo Scientific

Micro-Volume Sampling—How it Works

Surface Tension

Absorbance Spectrum

Sample Absorbance

Sample Concentration

Baseline Correction

Surface Tension

The NanoDrop One spectrophotometer uses surface tension to hold 
a small volume of sample between two pedestals. The patented 
sample retention system enables the measurement of highly 
concentrated samples without the need for dilutions.

A fiber optic cable embedded in the upper pedestal leads to a xenon 
light source. A second cable embedded in the lower pedestal leads to 
a detector. When the instrument arm is down, the sample forms a 
liquid column, essentially bridging the gap between the two fiber 
optic cables.

Summary of Contents for NanoDrop One

Page 1: ...NanoDrop Micro UV Vis Spectrophotometers NanoDrop One User Guide 269 309101 Revision B July 2016 ...

Page 2: ...her com Thermo Fisher Scientific Inc provides this document to its customers with a product purchase to use in the product operation This document is copyright protected and any reproduction of the whole or any part of this document is strictly prohibited except with the written authorization of Thermo Fisher Scientific Inc The contents of this document are subject to change without notice All tec...

Page 3: ...3 Measure Microarray Samples 23 Microarray Reported Results 27 Settings for Microarray Measurements 28 Calculations for Microarray Measurements 32 Measure using a Custom Factor 35 Measure Nucleic Acid using a Custom Factor 35 Custom Factor Reported Results 37 Settings for Nucleic Acid Measurements using a Custom Factor 39 Detection Limits for Nucleic Acid Measurements using a Custom Factor 39 Meas...

Page 4: ...Calculations for Protein A205 Measurements 85 Measure Protein BCA 87 Measure Total Protein Concentration 87 Protein BCA Reported Results 96 Settings for Protein BCA Measurements 100 Measure Protein Bradford 101 Measure Total Protein Concentration 101 Protein Bradford Reported Results 106 Settings for Protein Bradford Measurements 109 Measure Protein Lowry 111 Measure Total Protein Concentration 11...

Page 5: ...erations 210 NanoDrop One Home Screen 211 NanoDrop One Measurement Screens 215 NanoDrop One Data Viewer 222 NanoDrop One General Operations 229 Instrument Settings 236 Acclaro Sample Intelligence 240 NanoDrop One Viewer Software 248 Viewer Home Screen 249 Manage Experiments and Associated Data 251 Manage Identifiers on a PC 260 Manage Custom Methods 265 Multimedia 277 Chapter 4 Maintaining Your In...

Page 6: ...NanoDrop One User Guide Thermo Scientific Chapter 5 Safety and Operating Precautions 301 Operating Precautions 302 Safety Information 303 Chapter 6 About this Help System 311 Chapter 7 Contact Technical Support 313 ...

Page 7: ...loaded software and a touchscreen display The instrument can be connected to an optional USB label printer NOTICE Before operating a NanoDrop One instrument please read the safety and operating precautions and then follow their recommendations when using the instrument Instrument Models and Features There are two models available for the NanoDrop One spectrophotometer Register Your Instrument Regi...

Page 8: ... features The NanoDrop OneC model also features a cuvette holder for analyzing dilute samples using standard UV visible cuvettes Both instruments come with a built in 7 inch Android high resolution touchscreen preloaded with easy to use instrument control software The NanoDrop One software is loaded with features to integrate with and simplify your daily workflows 1Locate the instrument away from ...

Page 9: ...tures Thermo Scientific NanoDrop One User Guide 3 Touchscreen 1Two more USB A ports are located on instrument back panel Touchscreen can slide left or right to accommodate personal preference and tilt forward or back for optimal viewing USB A port1 Pedestals Touchscreen ...

Page 10: ...ons on demand technical support for measurements that are atypical or very low concentration invalid result alerts a column sensor monitors for the presence of bubbles or reflective particles that can compromise measurement results The NanoDrop OneC model includes a cuvette holder for measuring dilute samples colorimetric assays cell cultures and kinetic studies The cuvette system has these additi...

Page 11: ...s printing of data from each sample measurement or from a group of samples logged and measured together The printer connects to the instrument front or back via a USB cable included PR 1 Pedestal Reconditioning Kit PV 1 Performance Verification Solution Liquid photometric standard used to check instrument performance For more information see Performance Verification Specially formulated conditioni...

Page 12: ...ternet connection is required for registration To register your instrument 1 Do one of the following From the NanoDrop One Viewer software running on a personal computer PC that is connected to the Internet open the Help menu and choose NanoDrop One Website From any PC that is connected to the Internet use any web browser to navigate to our website 2 On the website locate NanoDrop One Registration...

Page 13: ...owing To install the Viewer software on a computer for the first time open any web browser and find the NanoDrop website To update or upgrade the Viewer software from the Viewer Home screen open the Help menu and choose NanoDrop One Website to open our website 2 On the NanoDrop website locate the software downloads page 3 Select to download NanoDrop One PC Viewer software English version and follo...

Page 14: ...rt the USB device into any USB port on the NanoDrop One instrument 6 From the instrument Home screen tap Settings System Update Software If the USB device contains more than one version of the installer a message is displayed Select the version to install English installer must be run first and tap Update An instrument restart is required after the English installer completes When the installation...

Page 15: ...e is 0 1 5 A Sample Type Lower Detection Limit Upper Detection Limit Typical Reproducibilitya dsDNA 2 0 ng μL pedestal 0 20 ng μL cuvette 27 500 ng μL pedestal 75 ng μL cuvette 2 0 ng μL for sample concentrations between 2 0 and 100 ng μL samples 2 for samples 100 ng μL ssDNA 1 3 ng μL pedestal 0 13 ng μL cuvette 18 150 ng μL pedestal 49 5 ng μL cuvette 2 0 ng μL for sample concentrations between ...

Page 16: ...g mL pedestal 2 over entire range Protein Bradford 100 μg mL 50 1 reagent sample volume 15 μg mL 1 1 reagent sample volume 8000 μg mL 100 μg μL 25 μg mL for 100 500 μg mL samples 5 for 500 8000 μg mL samples 4 μg mL for 15 50 μg mL samples 5 for 50 125 μg mL samples Protein Pierce 660 50 μg mL 15 1 reagent sample volume 25 μg mL 7 5 1 reagent sample volume 2000 μg mL 1000 μg mL 3 μg mL for 50 125 ...

Page 17: ... pmol μL for sample concentrations between 0 20 and 4 0 pmol μL 2 for samples 4 0 pmol μL Cy5 Cy5 5 Alexa Fluor 647 0 12 pmol μL pedestal 60 pmol μL pedestal 0 12 pmol μL for sample concentrations between 0 12 and 2 4 pmol μL 2 for samples 2 4 pmol μL Alexa Fluor 488 Alexa Fluor 594 0 4 pmol μL pedestal 215 pmol μL pedestal 0 40 pmol μL for sample concentrations between 0 40 and 8 0 pmol μL 2 for ...

Page 18: ......

Page 19: ...230 A single point baseline correction can also be used To measure dsDNA ssDNA or RNA samples Measures the concentration of purified dsDNA ssDNA or RNA samples that absorb at 260 nm Measure dsDNA ssDNA or RNA Reported Results Settings Detection Limits Calculations NOTICE Do not use a squirt or spray bottle on or near the instrument as liquids will flow into the instrument and may cause permanent d...

Page 20: ...t light path 4 Tap Blank and wait for the measurement to complete Tip If Auto Blank is On the blank measurement starts automatically after you lower the arm This option is not available for cuvette measurements 5 Lift the arm and clean both pedestals with a new laboratory wipe or remove the blanking cuvette 6 Pipette 1 2 μL sample solution onto the pedestal and lower the arm or insert the sample c...

Page 21: ...on See Choosing and Measuring a Blank for more information For micro volume measurements Ensure pedestal surfaces are properly cleaned and conditioned If possible heat highly concentrated or large molecule samples such as genomic or lambda DNA to 63 C 145 F and gently but thoroughly vortex before taking a measurement Avoid introducing bubbles when mixing and pipetting Follow best practices for mic...

Page 22: ... spectra Press and hold sample row to view measurement details Drag tab down up to see more less sample data Nucleic acid concentration UV spectrum Tap to select unit Purity ratios Menu of options tap to open Sample name tap to edit Swipe screen left to view table with more measurement results Pinch and zoom to adjust axes double tap to reset Tap to end experiment and export data Menu of options t...

Page 23: ...he sample row Here is an example sample details application and sampling method used i e pedestal or cuvette sample name created on date sample measurement was taken nucleic acid concentration A260 A280 A260 A230 A260 A280 factor baseline correction Related Topics Basic Instrument Operations Nucleic Acid Calculations Setting for Nucleic Acid Measurements To show the dsDNA ssDNA or RNA settings fro...

Page 24: ...absorbance of sample spectrum is zero at specified baseline correction wavelength The nucleic acid applications use the Beer Lambert equation to correlate absorbance with concentration Solving Beer s law for concentration yields the equation at the right Beer Lambert Equation solved for concentration c A b where A UV absorbance in absorbance units AU wavelength dependent molar absorptivity coeffic...

Page 25: ... molar extinction coefficient in ng cm μL b sample pathlength in cm As a result analyte concentration c is calculated as c A 1 b or c A f where c analyte concentration in ng μL A absorbance in absorbance units A f factor in ng cm μL see below For the dsDNA ssDNA and RNA applications the generally accepted factors for nucleic acids are used in conjunction with Beer s Law to calculate sample concent...

Page 26: ...er than 10 mm cuvettes the spectra are normalized to a 10 mm pathlength equivalent A260 absorbance Nucleic acid absorbance values are measured at 260 nm using the normalized spectrum This is the reported A260 value if Baseline Correction is not selected If Baseline Correction is selected the absorbance value at the correction wavelength is subtracted from the absorbance at 260 nm The corrected abs...

Page 27: ...m An A260 A280 purity ratio of 1 8 is generally accepted as pure for DNA 2 0 for RNA Acidic solutions may under represent the reported value by 0 2 0 3 the opposite is true for basic solutions A260 A230 purity ratio Ratio of corrected absorbance at 260 nm to corrected absorbance at 230 nm An A260 A230 purity ratio between 1 8 and 2 2 is generally accepted as pure for DNA and RNA Note Although puri...

Page 28: ......

Page 29: ...ntrations as low as 0 2 picomole per microliter To measure microarray samples Measures the concentration of purified nucleic acids that have been labeled with up to two fluorescent dyes for use in downstream microarray applications Measure Microarray Samples Reported Results Settings Detection Limits Calculations NOTICE Do not use a squirt or spray bottle on or near the instrument as liquids will ...

Page 30: ...ific Before you begin Before taking pedestal measurements with the NanoDrop One instrument lift the instrument arm and clean the upper and lower pedestals At a minimum wipe the pedestals with a new laboratory wipe For more information see Cleaning the Pedestals ...

Page 31: ...s automatically after you lower the arm This option is not available for cuvette measurements 5 Lift the arm and clean both pedestals with a new laboratory wipe or remove the blanking cuvette 6 Pipette 1 2 μL sample solution onto the pedestal and lower the arm or insert the sample cuvette into the cuvette holder 7 Start the sample measurement Pedestal If Auto Measure is On lower arm if Auto Measur...

Page 32: ...c Related Topics Best Practices for Nucleic Acid Measurements Measure a Micro Volume Sample Measure a Sample Using a Cuvette Best Practices for Micro Volume Measurements Best Practices for Cuvette Measurements Prepare Samples and Blanks Basic Instrument Operations ...

Page 33: ... and hold sample row to view measurement details Nucleic acid concentration Tap to select unit UV visible spectrum Menu of options tap to open Sample name tap to edit Swipe screen left to view table with more measurement results Pinch and zoom to adjust axes double tap to reset Tap to end experiment and export data Note A baseline correction is performed at 750 nm absorbance value at 750 nm is sub...

Page 34: ...estal or cuvette sample name created on date sample measurement was taken nucleic acid concentration A260 A260 A280 dye 1 dye 2 concentration sample type analysis correction factor Related Topics Basic Instrument Operations Microarray Calculations Settings for Microarray Measurements Microarray settings The Microarray Setup screen appears after you select the Microarray application from the Nuclei...

Page 35: ...ically based on widely accepted value for each base unit Custom with user specified factor in ng cm μL Enter factor between 15 ng cm μL and 150 ng cm μL Dye 1 Dye 2 Typea a To add a custom dye or edit the list of available dyes use the Dye Chromophore Editor Cy3 5 3 5 or 5 5 Alexa Fluor 488 546 555 594 647 or 660 Select pre defined dye s used to label sample material or one that has been added usi...

Page 36: ... Home screen tap Dye Chrom Editor from the Microarray or Proteins Labels measurement screen tap Settings Dye Chrom Editor These operations are available from the Dye Chromophore Editor Tap to add custom dye Tap to edit selected custom dye Tap to delete selected custom dye Locked dye pre defined cannot be edited or deleted Selected dyes will appear in Dye1 and Dye2 lists in Microarray Setup or Prot...

Page 37: ...l be used to measure dye s absorbance specify dye s correction values at 260 nm and 280 nm tap Add Dye When a custom dye is selected before a measurement the dye s absorbance and concentration values are reported and the corrections are applied to the measured sample absorbance values and to the resulting sample concentrations and purity ratios Edit custom dye tap to select custom dye tap Note To ...

Page 38: ...application offers six options shown at right for selecting an appropriate factor for each measured sample to be used in conjunction with Beer s Law to calculate sample concentration If the factor is known choose the Custom Factor option and enter the factor in ng cm μL Otherwise choose the option that best matches the sample solution Tip Ideally the factor or extinction coefficient should be dete...

Page 39: ...lected the absorbance value at the correction wavelength is subtracted from the absorbance at 260 nm If one or more dyes are selected the dye correction values at 260 nm are also subtracted from the absorbance at 260 nm The final corrected absorbance at 260 nm is reported and used to calculate sample concentration A280 absorbance 750 corrected and normalized absorbance value at 280 nm minus the A2...

Page 40: ... selected unit i e ng μL μg uL or μg mL Calculations are based on modified Beer s Law equation using corrected nucleic acid absorbance value A260 A280 purity ratio Ratio of corrected absorbance at 260 nm to corrected absorbance at 280 nm An A260 A280 purity ratio of 1 8 is generally accepted as pure for DNA 2 0 for RNA Acidic solutions may under represent the reported value by 0 2 0 3 the opposite...

Page 41: ...nd A260 A230 A single point baseline correction can also be used To measure nucleic acid samples using a custom factor Measures the concentration of purified nucleic acids using a custom factor for the calculations Measure using Custom Factor Reported Results Settings Detection Limits Calculations NOTICE Do not use a squirt or spray bottle on or near the instrument as liquids will flow into the in...

Page 42: ... align the cuvette light path with the instrument light path 4 Tap Blank and wait for the measurement to complete Tip If Auto Blank is On the blank measurement starts automatically after you lower the arm This option is not available for cuvette measurements 5 Lift the arm and clean both pedestals with a new laboratory wipe or remove the blanking cuvette 6 Pipette 1 2 μL sample solution onto the p...

Page 43: ...ette Measurements Prepare Samples and Blanks Basic Instrument Operations Custom Factor Reported Results For each measured sample this application shows the absorbance spectrum and a summary of the results Here is an example Note The Custom Factor measurement screen is identical to the measurement screen for the other nucleic acid applications except the Custom Factor is reported in the lower left ...

Page 44: ... Custom Factor 38 NanoDrop One User Guide Thermo Scientific Related Topics Basic Instrument Operations Nucleic Acid Reported Results Nucleic Acid Calculations Custom factor used to calculate nucleic acid concentration ...

Page 45: ... extinction coefficientsample Setting Available Options Description Custom Factor Enter an integer value between 15 ng cm μL and 150 ng cm μL Constant used to calculate nucleic acid concentration in modified Beer s Law equation Based on extinction coefficient and pathlength f 1 260 b where f factor molar extinction coefficient at 260 nm in ng cm μL b sample pathlength in cm 1 cm for nucleic acids ...

Page 46: ...le measurement using an extinction coefficient of 55 the equation looks like this 550 AU 55 ng cm μL 30 250 ng μL Related Topics Detection Limits for All Applications Note For measurements with 10 mm pathlength cuvettes the upper absorbance limit is 1 5 AU which is approximately 75 ng μL for dsDNA ...

Page 47: ...point baseline correction can also be used Measures the concentration of purified ssDNA or RNA oligonucleotides that absorb at 260 nm Measure Oligo DNA or RNA Reported Results Settings Detection Limits Calculations Note If the oligonucleotide has been modified for example with a fluorophore dye check with the oligo manufacturer to determine if the modification contributes absorbance at 260 nm If i...

Page 48: ...strument arm and clean the upper and lower pedestals At a minimum wipe the pedestals with a new laboratory wipe For more information see Cleaning the Pedestals NOTICE Do not use a squirt or spray bottle on or near the instrument as liquids will flow into the instrument and may cause permanent damage Do not use hydrofluoric acid HF on the pedestals Fluoride ions will permanently damage the quartz f...

Page 49: ...urement to complete Tip If Auto Blank is On the blank measurement starts automatically after you lower the arm This option is not available for cuvette measurements 5 Lift the arm and clean both pedestals with a new laboratory wipe or remove the blanking cuvette 6 Pipette 1 2 μL sample solution onto the pedestal and lower the arm or insert the sample cuvette into the cuvette holder 7 Start the sam...

Page 50: ...Drop One User Guide Thermo Scientific Measure a Micro Volume Sample Measure a Sample Using a Cuvette Best Practices for Micro Volume Measurements Best Practices for Cuvette Measurements Prepare Samples and Blanks Basic Instrument Operations ...

Page 51: ...lect sample and update spectrum tap more rows to overlay up to five spectra Press and hold sample row to view measurement details Drag tab down up to see more less sample data UV spectrum1 Purity ratios Tap to select unit Sample name tap to edit Menu of options tap to open Swipe screen left to view table with more measurement results Pinch and zoom to adjust axes double tap to reset Tap to end exp...

Page 52: ... sample row Here is an example sample details application and sampling method used i e pedestal or cuvette sample name created on date sample measurement was taken nucleic acid concentration A260 A280 A260 A230 A260 A280 factor oligo sequence baseline correction stirrer status Related Topics Basic Instrument Operations Oligo Calculations Note The five nucleotides that comprise DNA and RNA exhibit ...

Page 53: ... C keys to specify the DNA base sequence for RNA Use the G A U and C keys to specify the RNA base sequence Specify your DNA or RNA base sequence by tapping the corresponding keys Each time a base is added to the sequence the software calculates the following Factor Constant used to calculate oligonucleotide concentration in modified Beer s Law equation Based on extinction coefficient and pathlengt...

Page 54: ... Weight of oligo calculated from user defined base sequence Number of Bases entered Molar Ext Coefficient 260 nm Molar extinction coefficient of oligo in ng cm μL at 260 nm calculated from entered base sequence GC Percentage of guanine and cytosine residues in total number of bases entered Baseline Correction On or off Enter baseline correction wavelength in nm or use default value 340 nm Corrects...

Page 55: ...r natural essentially randomized sequences but not for short synthetic oligo sequences To ensure the most accurate results we use the exact value of 260 to calculate oligonucleotide concentration The NanoDrop software allows you to specify the base sequence of an oligonucleotide before it is measured For any entered base sequence the software uses the equation at the right to calculate the extinct...

Page 56: ...ents taken with nonstandard other than 10 mm cuvettes the spectra are normalized to a 10 mm pathlength equivalent Nucleic acid absorbance values are measured at 260 nm using the normalized spectrum This is the reported A260 value if Baseline Correction is not selected If Baseline Correction is selected the absorbance value at the correction wavelength is subtracted from the sample absorbance at 26...

Page 57: ...her 260 280 ratio due to the higher ratio of Uracil compared to that of Thymine Reported Values Nucleic acid concentration Reported in selected unit i e ng μL μg uL or μg mL Calculations are based on modified Beer s Law equation using corrected nucleic acid absorbance value A260 A280 purity ratio Ratio of corrected absorbance at 260 nm to corrected absorbance at 280 nm A260 A230 purity ratio Ratio...

Page 58: ......

Page 59: ...be used This application does not require a standard curve To measure Protein A280 samples Measures the concentration of purified protein populations that absorb at 280 nm Measure A280 Proteins Reported Results Settings Detection Limits Calculations Note If your samples contain mainly peptide bonds and little or no amino acids use the Protein A205 application instead of Protein A280 NOTICE Do not ...

Page 60: ...tific Before you begin Before taking pedestal measurements with the NanoDrop One instrument lift the instrument arm and clean the upper and lower pedestals At a minimum wipe the pedestals with a new laboratory wipe For more information see Cleaning the Pedestals ...

Page 61: ...utomatically after you lower the arm This option is not available for cuvette measurements 5 Lift the arm and clean both pedestals with a new laboratory wipe or remove the blanking cuvette 6 Pipette 2 μL sample solution onto the pedestal and lower the arm or insert the sample cuvette into the cuvette holder 7 Start the sample measurement Pedestal If Auto Measure is On lower arm if Auto Measure is ...

Page 62: ...rves Run a blanking cycle to assess the absorbance contribution of your buffer solution If the buffer exhibits strong absorbance at or near the analysis wavelength typically 280 nm or 205 nm you may need to choose a different buffer or application such as a colorimetric assay for example BCA or Pierce 660 See Choosing and Measuring a Blank for more information For micro volume measurements Ensure ...

Page 63: ...easurements Measure a Micro Volume Sample Measure a Sample Using a Cuvette Prepare Samples and Blanks Basic Instrument Operations Protein A280 Reported Results Protein A280 measurement screen For each measured sample this application shows the absorbance spectrum and a summary of the results Here is an example ...

Page 64: ...lay up to five spectra Press and hold sample row to view measurement details Drag tab down up to see more less sample data Swipe screen left to view table with more measurement results Pinch and zoom to adjust axes double tap to reset Tap to end experiment and export data UV spectrum Tap to select unit Menu of options tap to open Sample name tap to edit Protein concentration Absorbance at 280 nm P...

Page 65: ...n A280 settings from the Protein A280 measurement screen tap Protein A280 Setup Protein A280 settings The Protein A280 application provides a variety of sample type options for purified protein analysis Sample name tap to edit Sampling method Application Date time measured Protein conc Absorbance at 280 nm Sample type Baseline correction wavelength Baseline correction absorbance Purity ratio ...

Page 66: ... rough estimate of protein concentration is acceptable for a solution with no other interfering substances Assumes 0 1 1 mg mL protein solution produces 1 0A at 280 nm where pathlength is 10 mm i e 1 10 BSA 6 67 Calculates BSA Bovine Serum Albumin protein concentration using mass extinction coefficient of 6 67 L gm cm at 280 nm for 1 i e 10 mg mL BSA solution Assuming MW is 66 400 daltons Da molar...

Page 67: ...r extinction coefficient of 210 000 M 1cm 1 enter 210 Other protein 1 User entered mass extinction coefficient Assumes protein has known mass extinction coefficient Enter mass extinction coefficient in L gm cm for 10 mg mL 1 protein solution Baseline Correction On or off Enter baseline correction wavelength in nm or use default value 340 nm N A Corrects for any offset caused by light scattering pa...

Page 68: ...r Description for new protein specify whether to enter Molar Extinction coefficient or Mass Extinction coefficient for custom protein if Mass Extinction coefficient is selected enter mass extinction coefficient in L gm cm for 10 mg mL 1 protein solution Tap to add custom protein Tap to edit selected custom protein Tap to delete selected custom protein Custom proteins will appear in Sample Type lis...

Page 69: ...icient of 210 000 M 1cm 1 enter 210 enter molecular weight MW in kiloDaltons kDa tap OK to close New Protein Type box After you choose OK the new custom protein appears in the Type list in Protein A280 Setup and Proteins Labels Setup Edit custom protein in Protein Editor tap to select custom protein tap to show Edit Protein Type box edit any entries or settings tap OK Tap a field to show keyboard ...

Page 70: ...endent on the upper absorbance limit of the instrument and the sample s extinction coefficient To calculate upper detection limits for other non BSA protein sample types To calculate upper detection limits in ng μL for proteins use the following equation upper absorbance limitinstrument mass extinction coefficientsample 10 For example if the sample s mass extinction coefficient at 280 nm is 6 67 f...

Page 71: ...s molar extinction coefficient yields the molar concentration of the sample See Published Extinction Coefficients for more information regarding molar vs mass concentration values The extinction coefficient of a peptide or protein is related to its tryptophan W tyrosin Y and cysteine C amino acid composition Tip The extinction coefficient is wavelength specific for each protein and can be affected...

Page 72: ...fied mass ext coefficient Note See Sample Type for details Most sources report extinction coefficients for proteins measured at or near 280 nm in phosphate or other physiologic buffer These values provide sufficient accuracy for routine assessments of protein concentration Published Extinction Coefficients Published extinction coefficients for proteins may be reported as wavelength dependent molar...

Page 73: ...otein samples Purity ratios are sensitive to the presence of contaminants in the sample such as residual solvents and reagents typically used during sample purification Measured Values A280 absorbance Note For micro volume absorbance measurements and measurements taken with nonstandard other than 10 mm cuvettes the spectra are normalized to a 10 mm pathlength equivalent Protein absorbance values a...

Page 74: ...ng corrected protein absorbance value A260 A280 purity ratio Ratio of corrected absorbance at 260 nm to corrected absorbance at 280 nm An A260 A280 purity ratio of 0 57 is generally accepted as pure for proteins Note Although purity ratios are important indicators of sample quality the best indicator of protein quality is functionality in the downstream application of interest e g real time PCR ...

Page 75: ...ction of dye concentrations as low as 0 2 picomole per microliter This information is useful for evaluating protein dye conjugation degree of labeling for use in downstream applications To measure labeled protein samples Measures the concentration of purified proteins that have been labeled with up to two fluorescent dyes Measure Labeled Proteins Reported Results Settings Detection Limits Calculat...

Page 76: ...ientific Before you begin Before taking pedestal measurements with the NanoDrop One instrument lift the instrument arm and clean the upper and lower pedestals At a minimum wipe the pedestals with a new laboratory wipe For more information see Cleaning the Pedestals ...

Page 77: ...fter you lower the arm This option is not available for cuvette measurements 5 Lift the arm and clean both pedestals with a new laboratory wipe or remove the blanking cuvette 6 Pipette 2 μL sample solution onto the pedestal and lower the arm or insert the sample cuvette into the cuvette holder 7 Start the sample measurement Pedestal If Auto Measure is On lower arm if Auto Measure is off lower arm ...

Page 78: ...easurements Measure a Micro Volume Sample Measure a Sample Using a Cuvette Prepare Samples and Blanks Basic Instrument Operations Proteins Labels Reported Results Proteins Labels measurement screen For each measured sample this application shows the absorbance spectrum and a summary of the results Here is an example ...

Page 79: ...s tap to open Sample name tap to edit Protein concentration Dye concentration s Swipe screen left to view table with more measurement results Pinch and zoom to adjust axes double tap to reset Tap to end experiment and export data Note A baseline correction is performed at 750 nm absorbance value at 750 nm is subtracted from absorbance values at all wavelengths in sample spectrum Micro volume absor...

Page 80: ...w Here is an example Reported values for Proteins Labels application Sample details application and sampling method used i e pedestal or cuvette Sample Name Creation date Protein A280 Sample Type Dye 1 Dye 2 Sloping Dye Correction Analysis Correction Related Topics Basic Instrument Operations Proteins Labels calculations Settings for Proteins and Labels Measurements To show the Proteins Labels set...

Page 81: ...cts calculation for protein concentration only On or off Enter analysis correction wavelength in nm or use default value 340 nm N A Corrects sample absorbance measurement for any offset caused by light scattering particulates by subtracting absorbance value at specified analysis correction wavelength from absorbance value at analysis wavelength Corrected value is used to calculate sample concentra...

Page 82: ... apply to any protein sample type The upper detection limits are dependent on the upper absorbance limit of the instrument and the sample s extinction coefficient To calculate upper detection limits for other non BSA protein sample types To calculate upper detection limits in ng μL for proteins use the following equation upper absorbance limitinstrument mass extinction coefficientsample 10 For exa...

Page 83: ...ficient Other protein 1 user specified mass ext coefficient Note See Sample Type for details Calculated protein concentrations are based on the absorbance value at 280 nm the selected or entered extinction coefficient and the sample pathlength A single point baseline correction or analysis correction may be applied Concentration is reported in mass units Calculators are available on the Internet t...

Page 84: ... calculate dye concentrations Dye correction Pre defined dyes have known correction values for A260 and A280 See Dye Chromophore Editor for correction values used A280 dye correction is subtracted from A280 absorbance value used to calculate protein concentration Sample Pathlength For micro volume measurements the software selects the optimal pathlength between 1 0 mm and 0 03 mm based on sample a...

Page 85: ...tandard curve To measure Protein A205 samples Measures the concentration of purified protein populations that absorb at 205 nm Measure A205 Proteins Reported Results Settings Detection Limits Calculations Note If your samples contain mainly amino acids such as tryptophan or tyrosine or cys cys disulfide bonds use the Protein A280 application instead of Protein A205 NOTICE Do not use a squirt or sp...

Page 86: ... the cuvette holder Tip If using a cuvette make sure to align the cuvette light path with the instrument light path 4 Tap Blank and wait for the measurement to complete Tip If Auto Blank is On the blank measurement starts automatically after you lower the arm This option is not available for cuvette measurements 5 Lift the arm and clean both pedestals with a new laboratory wipe or remove the blank...

Page 87: ...Measure Protein A205 Thermo Scientific NanoDrop One User Guide 81 Prepare Samples and Blanks Basic Instrument Operations ...

Page 88: ... Press and hold sample row to view measurement details Drag tab down up to see more less sample data Swipe screen left to view table with more measurement results Pinch and zoom to adjust axes double tap to reset Tap to end experiment and export data UV spectrum Tap to select unit Menu of options tap to open Sample name tap to edit Protein concentration Absorbance at 205 nm Absorbance at 280 nm No...

Page 89: ...d hold the sample row Here is an example Related Topics Basic Instrument Operations Protein A205 Calculations Settings for Protein A205 Measurements To show the Protein A205 settings from the Protein A205 measurement screen tap Protein A205 Setup Sample name tap to edit Sampling method Application Date time measured Protein conc Absorbance at 205 nm Sample type Baseline Correction wavelength Basel...

Page 90: ...ss extinction coefficient in L gm cm for 1 mg mL 0 1 protein solution Baseline Correction On or off Enter baseline correction wavelength in nm or use default value 340 nm N A Corrects for any offset caused by light scattering particulates by subtracting measured absorbance at specified baseline correction wavelength from absorbance values at all wavelengths in sample spectrum As a result absorbanc...

Page 91: ...m 27 120 A280 A205 Other protein enter mass extinction coefficient in L gm cm for 1 mg mL 0 1 protein solution Note See Sample Type for details Calculated protein concentrations are based on the absorbance value at 205 nm the selected or entered extinction coefficient and the sample pathlength A single point baseline correction may also be applied Concentration is reported in mass units Calculator...

Page 92: ...d 0 03 mm based on sample absorbance at the analysis wavelength For cuvette measurements pathlength is determined by the cuvette Pathlength setting in the software see General Settings Displayed spectra and absorbance values are normalized to a 10 mm pathlength equivalent Reported Values Protein concentration Reported in selected unit mg mL or μg mL Calculations are based on Beer Lambert equation ...

Page 93: ...concentration A single point baseline correction is applied Theory of Protein BCA assay The Protein BCA assay uses bicinchoninic acid BCA as the detection reagent for Cu 1 which is formed when Cu 2 is reduced by certain proteins in an alkaline environment A purple reaction product is formed by the chelation of two molecules of BCA with one cuprous ion Cu 1 The resulting Cu BCA chelate formed in th...

Page 94: ...way See the kit manufacturer s guidelines and recommendations All reference and standards solutions should be the same buffer used to resuspend the samples plus the same volume of reagent added to the samples First standard is a reference measurement The reference solution should contain none of the analyte of interest The reference measurement is not the same as a blank measurement This applicati...

Page 95: ...rence and all standards before you start analyzing samples After the first sample has been measured no additional changes are allowed to the standard curve As you measure the standards a measurement screen appears similar to the measurement screens for samples Reference concentration and absorbance value Standard concentrations and absorbance values Swipe left one screen to view standard curve Men...

Page 96: ...e The R2 value indicates how well the standard curve fits the standard data points 1 0 is a perfect fit all points lie exactly on the curve Swipe left one screen to view data table for standards Curve type setting R2 value 1 0 equals perfect fit White circles indicate data points for standards Standard curve Press and hold any row to view details ...

Page 97: ...Swipe left one screen to see the data table for the standards Here is an example Press and hold a row in any of the previous screens to view details about an individual standard Here is an example Press and hold any row to view details Tap to delete this measurement ...

Page 98: ...rd from standards measurement screen tap application name Setup tap the next empty Concentration field and enter the concentration value for the new standard tap Done Edit standard from standards measurement screen tap application name Setup tap the Concentration field and edit the concentration value tap Done Delete standard from standards measurement screen standard curve screen or standards dat...

Page 99: ...Protein BCA standards and samples Before you begin Before taking pedestal measurements with the NanoDrop One instrument lift the instrument arm and clean the upper and lower pedestals At a minimum wipe the pedestals with a new laboratory wipe For more information see Cleaning the Pedestals NOTICE Do not use a squirt or spray bottle on or near the instrument as liquids will flow into the instrument...

Page 100: ...t measurement 5 Measure remaining standards pipette 2 μL standard 1 onto pedestal or insert standard 1 cuvette lower arm to start measurement or tap Measure if Auto Measure is off lift arm and clean both pedestals with new wipe or remove cuvette if Replicates setting is greater than 1 repeat measurement repeat substeps above for each additional standard when specified number of standards and repli...

Page 101: ...Measure Protein BCA Thermo Scientific NanoDrop One User Guide 95 Prepare Samples and Blanks Basic Instrument Operations ...

Page 102: ...ndard curve is also available by swiping left from the measurement screen or in the Data Viewer as shown below Tap row to select sample and update spectrum tap more rows to overlay up to five spectra Press and hold sample row to view measurement details Drag tab down up to see more less sample data Swipe screen left to view standard curve Pinch and zoom to adjust axes double tap to reset Visible s...

Page 103: ...r a selected sample A horizontal line connects the sample absorbance value on the Y axis to the standard curve A vertical line connects that point to the sample concentration value on the X axis Note A baseline correction is performed at 750 nm absorbance value at 750 nm is subtracted from absorbance values at all wavelengths in sample spectrum Micro volume absorbance measurements and measurements...

Page 104: ...es for up to five samples Press and hold sample row for measurement details Drag tab down up to see more less sample data Pinch and zoom to adjust axes double tap to reset Standard curve Menu of options tap to open Protein concentration for selected sample Absorbance at 562 nm Page control swipe screen left or right to view next or previous screen Curve type R2 value 1 0 equals perfect fit Red squ...

Page 105: ...s screen see previous image show a summary of the reported values To view all reported values press and hold the sample row Here is an example Related Topics Basic Instrument Operations Protein A280 Calculations Sample name tap to edit Sampling method Date time measured Protein conc Absorbance at 562 nm Baseline correction wavelength Baseline correction absorbance ...

Page 106: ...ndards requires reference measurement and at least one standard Interpolation Draws a series of straight lines to connect all measured standards requires reference measurement and at least one standard 2nd order polynomial Draws the 2nd order least squares polynomial using all measured standards requires reference measurement and at least two standards 3rd order polynomial Draws the 3rd order leas...

Page 107: ...595 nm and uses a standard curve to calculate protein concentration See Working with Standard Curves for more information A single point baseline correction is applied Theory of Protein Bradford assay The Protein Bradford assay uses the protein induced absorbance shift of Coomassie Blue dye to determine total protein concentration The bound protein dye complex is measured at 595 nm and baseline co...

Page 108: ...ations for all standards and samples unknowns Ensure each is subjected to the same timing and temperature throughout the assay Protein standards for generating a standard curve may also be provided by the kit manufacturer Since the NanoDrop One pedestals can measure higher protein concentrations than traditional cuvette based spectrophotometers you may need to supply your own protein standards at ...

Page 109: ...For more information see Cleaning the Pedestals To measure Protein Bradford standards and samples 1 From the Home screen select the Proteins tab and tap Protein Bradford 2 Specify a curve type and number of replicates for each standard and enter the concentration of each standard Tip For this assay set Curve Type to 2nd Order Polynomial and Replicates to 3 3 Measure blank pipette 2 μL DI H2O onto ...

Page 110: ...at measurement 5 Measure remaining standards pipette 2 μL standard 1 onto pedestal or insert standard 1 cuvette lower arm to start measurement or tap Measure if Auto Measure is off lift arm and clean both pedestals with new wipe or remove cuvette if Replicates setting is greater than 1 repeat measurement repeat substeps above for each additional standard when specified number of standards and repl...

Page 111: ...Measure Protein Bradford Thermo Scientific NanoDrop One User Guide 105 Measure a Sample Using a Cuvette Prepare Samples and Blanks Basic Instrument Operations ...

Page 112: ... The standard curve is also available by swiping left from the measurement screen or in the Data Viewer as shown below Tap row to select sample and update spectrum tap more rows to overlay up to five spectra Press and hold sample row to view measurement details Drag tab down up to see more less sample data Swipe screen left to view standard curve Pinch and zoom to adjust axes double tap to reset V...

Page 113: ... value on the Y axis to the standard curve A vertical line connects that point to the sample concentration value on the X axis The R2 value indicates how well the standard curve fits the standard data points 1 0 is a perfect fit that is all points lie exactly on the curve Note A baseline correction is performed at 750 nm absorbance value at 750 nm is subtracted from absorbance values at all wavele...

Page 114: ...p to see more less sample data Pinch and zoom to adjust axes double tap to reset Standard curve Menu of options tap to open Protein concentration for selected sample Absorbance at 595 nm Page control swipe screen left or right to view next or previous screen Curve type R2 value 1 0 equals perfect fit Red squares indicate sample data points White circles indicate standard data points Selected sampl...

Page 115: ... screen tap Protein Bradford Setup Sample name tap to edit Sampling method Date time measured Protein conc Absorbance at 595 nm Baseline correction wavelength Baseline correction absorbance Note You can edit the Curve Type setting when measuring standards by changing the list box at the top of the application measurement screen You can edit the concentration value for a standard from the applicati...

Page 116: ... polynomial Draws the 2nd order least squares polynomial using all measured standards requires reference measurement and at least two standards 3rd order polynomial Draws the 3rd order least squares polynomial using all measured standards requires reference measurement and at least three standards Replicates Enter number of measurements of the reference or the same standard or sample that are aver...

Page 117: ... with Standard Curves for more information A single point baseline correction is applied Theory of Protein Lowry assay The Protein Lowry assay involves the reaction of protein with cupric sulfate in alkaline solution resulting in the formation of tetradentate copper protein complexes The Folin Ciocalteu reagent is effectively reduced in proportion to the chelated copper complexes The water soluble...

Page 118: ...For more information see Cleaning the Pedestals NOTICE Do not use a squirt or spray bottle on or near the instrument as liquids will flow into the instrument and may cause permanent damage Do not use hydrofluoric acid HF on the pedestals Fluoride ions will permanently damage the quartz fiber optic cables To measure Protein Lowry standards and samples 1 From the Home screen select the Proteins tab ...

Page 119: ...t measurement 5 Measure remaining standards pipette 2 μL standard 1 onto pedestal or insert standard 1 cuvette lower arm to start measurement or tap Measure if Auto Measure is off lift arm and clean both pedestals with new wipe or remove cuvette if Replicates setting is greater than 1 repeat measurement repeat substeps above for each additional standard when specified number of standards and repli...

Page 120: ...ce spectrum and a summary of the results The standard curve is also available by swiping left from the measurement screen or in the Data Viewer as shown below Tap row to select sample and update spectrum tap more rows to overlay up to five spectra Press and hold sample row to view measurement details Drag tab down up to see more less sample data Swipe screen left to view standard curve Pinch and z...

Page 121: ...lue on the Y axis to the standard curve A vertical line connects that point to the sample concentration value on the X axis The R2 value indicates how well the standard curve fits the standard data points 1 0 is a perfect fit that is all points lie exactly on the curve Note A baseline correction is performed at 405 nm absorbance value at 405 nm is subtracted from absorbance values at all wavelengt...

Page 122: ...ils Drag tab down up to see more less sample data Pinch and zoom to adjust axes double tap to reset Standard curve Menu of options tap to open Protein concentration for selected sample Absorbance at 650 nm Page control swipe screen left or right to view next or previous screen Curve type R2 value 1 0 equals perfect fit Red squares indicate sample data points White circles indicate standard data po...

Page 123: ...dards screen see previous image show a summary of the reported values To view all reported values press and hold the sample row Here is an example Related Topics Example standard curve Basic Instrument Operations Sample name tap to edit Sampling method Date time measured Protein conc Absorbance at 650 nm Baseline correction wavelength Baseline correction absorbance ...

Page 124: ...d standards requires reference measurement and at least one standard Interpolation Draws a series of straight lines to connect all measured standards requires reference measurement and at least one standard 2nd order polynomial Draws the 2nd order least squares polynomial using all measured standards requires reference measurement and at least two standards 3rd order polynomial Draws the 3rd order...

Page 125: ...660 assay The Protein Pierce 660 assay is based on the binding of a proprietary dye metal complex to protein in acidic conditions that causes a shift in the dye s absorption maximum which is measured at 660 nm The dye metal complex is reddish brown and changes to green upon protein binding The color change is produced by deprotonation of the dye at low pH facilitated by interactions with positivel...

Page 126: ...ame timing and temperature throughout the assay Protein standards for generating a standard curve may also be provided by the kit manufacturer Since the NanoDrop One pedestals can measure higher protein concentrations than traditional cuvette based spectrophotometers you may need to supply your own protein standards at higher concentrations than provided by the manufacturer For example additional ...

Page 127: ...rve type and number of replicates for each standard and enter the concentration of each standard Tip For this assay we recommend setting Curve Type to Linear 3 Measure blank Pipette 2 μL reference solution onto lower pedestal and lower arm or insert reference solution blanking cuvette into cuvette holder reference solution should contain none of the standard protein stock see Working With Standard...

Page 128: ...eat measurement 5 Measure remaining standards pipette 2 μL standard 1 onto pedestal or insert standard 1 cuvette lower arm to start measurement or tap Measure if Auto Measure is off lift arm and clean both pedestals with new wipe or remove cuvette if Replicates setting is greater than 1 repeat measurement repeat substeps above for each additional standard when specified number of standards and rep...

Page 129: ...Measure Protein Pierce 660 Thermo Scientific NanoDrop One User Guide 123 Measure a Sample Using a Cuvette Prepare Samples and Blanks Basic Instrument Operations ...

Page 130: ...le and update spectrum tap more rows to overlay up to five spectra Press and hold sample row to view measurement details Drag tab down up to see more less sample data Swipe screen left to view standard curve Pinch and zoom to adjust axes double tap to reset Visible spectrum Menu of options tap to open Protein concentration Absorbance at 660 nm Note A baseline correction is performed at 750 nm abso...

Page 131: ...indicates how well the standard curve fits the standard data points 1 0 is a perfect fit that is all points lie exactly on the curve Tap row to select sample and update graph tap more rows to overlay abs conc lines for up to five samples Press and hold sample row for measurement details Drag tab down up to see more less sample data Pinch and zoom to adjust axes double tap to reset Standard curve M...

Page 132: ... standards screen see previous image show a summary of the reported values To view all reported values press and hold the sample row Here is an example Related Topics Example standard curve Basic Instrument Operations Sample name tap to edit Sampling method Date time measured Protein conc Absorbance at 660 nm Baseline correction wavelength Baseline correction absorbance ...

Page 133: ...n Draws a series of straight lines to connect all measured standards requires reference measurement and at least one standard 2nd order polynomial Draws the 2nd order least squares polynomial using all measured standards requires reference measurement and at least two standards 3rd order polynomial Draws the 3rd order least squares polynomial using all measured standards requires reference measure...

Page 134: ...Measure Protein Pierce 660 128 NanoDrop One User Guide Thermo Scientific Instrument Settings ...

Page 135: ...l and stationary The OD600 application reports cell concentration in cells mL A single point absorbance correction can be used This application does not require a standard curve Theory of OD600 application The OD600 application measures light transmission and uses that value to calculate absorbance In spectroscopy transmitted light is defined as any light that is not absorbed by reflected from and...

Page 136: ... the same type of spectrophotometer and method i e pedestal vs cuvette as the amount of scattered light captured varies based on the optical configuration When using a different spectrophotometer or method calculate and apply a conversion factor to the reported results For example to compare OD readings using the pedestal vs a cuvette a conversion factor can be calculated as follows Conversion fac...

Page 137: ...ce at 600 nm use an alternative wavelength such as 400 nm to measure absorbance or use cuvettes instead of micro volume measurements For cuvette measurements NanoDrop OneC instruments only Use clean plastic glass or quartz cuvettes Follow best practices for cuvette measurements Do not use the automatic stirring feature for this assay To measure OD600 samples Before you begin Before taking pedestal...

Page 138: ...ath with the instrument light path 4 Tap Blank and wait for the measurement to complete Tip If Auto Blank is On the blank measurement starts automatically after you lower the arm This option is not available for cuvette measurements 5 Lift the arm and clean both pedestals with a new laboratory wipe or remove the blanking cuvette 6 Pipette 2 μL sample solution onto the pedestal and lower the arm or...

Page 139: ...ve spectra Press and hold sample row to view measurement details Drag tab down up to see more less sample data Swipe screen left to view table with more measurement results Pinch and zoom to adjust axes double tap to reset UV visible spectrum Menu of options tap to open Correctedabsorbance at 600 nm Corrected absorbance at additional wavelength Absorbance correction Additional wavelength Note Micr...

Page 140: ...d the sample row Here is an example Related Topics Basic Instrument Operations OD600 Calculations Settings for OD600 Measurements To show the OD600 settings from the OD600 measurement screen tap OD600 Setup Sample name tap to edit Sampling method Application Date time measured Corrected abs at 600 nm Absorbance correction Factor Cell culture concentration A600 Factor Additional wavelength Correcte...

Page 141: ...or displayed spectrum This can be useful for example to correct baseline offset caused by any difference between the media solution used to blank the instrument and media used to suspend the cell culture sample and because scattered light generally produces an offset Absorbance correction value is subtracted from absorbance values at all wavelengths in sample spectrum All displayed absorbance valu...

Page 142: ...ar absorption coefficient or extinction coefficient at specified wavelength b pathlength in cm 1 0 cm for the NanoDrop One instruments Cell number conversion factor 108 Any number User defined factor Generally accepted factor for measured cell type or one derived empirically using a solution of study cells at known concentration using the same media Default value is 1x108 which is the generally ac...

Page 143: ...rbance Correction is specified this is the reported A600 value and the value used to calculate cell concentration If an Absorbance Correction is specified the normalized and absorbance corrected absorbance value at 600 nm is reported and used to calculate cell concentration A absorbance Normalized and absorbance corrected if used absorbance value at any specified Additional Monitored Wavelength is...

Page 144: ......

Page 145: ...the instrument the method must also reside on the instrument This is the only way to run a custom method if your instrument is not connected to the computer with an Ethernet cable or through a wireless network Runs a custom measurement method created using NanoDrop One Viewer software Measure Custom Method Delete Custom Method Reported Results Note If the computer is connected to the instrument wi...

Page 146: ...hod files have a method filename extension 2 Connect the USB device to one of the USB ports on the instrument 3 From the Home screen select the Custom tab and tap Custom 4 Use the list box at the top of the screen to indicate the USB port used 5 Tap Load Method A message box shows the NanoDrop One methods available on the selected USB device 6 Tap one or more method names in the Load Method box to...

Page 147: ...equently acquired measurement results in the database on the instrument set Data Storage to Local see example above To run a custom method that resides on a personal computer connected to the instrument with an Ethernet cable and store all subsequently acquired measurement results in the database on that computer set Data Storage to Direct Connect PC see Set Up Ethernet Connection in Set Up the In...

Page 148: ... Method for more information If Data Storage is set to Direct Connect PC Ethernet or a specific computer name wireless Custom Setup shows only custom methods that reside on the wired Ethernet or specified wireless computer see Create Custom Method for more information 3 In the Select Method box tap to select the method to run Information about the selected method appears in the Method Details box ...

Page 149: ...ated Topics Measure a Micro Volume Sample Measure a Sample Using a Cuvette Create Custom Method Set Up the Instrument Export Custom Method Delete Custom Method From Home screen select Custom tab and tap Custom In Select Method box tap to select method to delete tap ...

Page 150: ...ple and update spectrum tap more rows to overlay up to five spectra Press and hold sample row to view measurement details Drag tab down up to see more less sample data UV visible spectrum Menu of options tap to open Analyte concentration Swipe screen left to view table with more measurement results Pinch and zoom to adjust axes double tap to reset Note Micro volume absorbance measurements and meas...

Page 151: ...appears after each measurement see previous image shows a summary of the reported values To view all reported values press and hold the sample row Here is an example Related Topics Basic Instrument Operations Sample name tap to edit Sampling method Method name Date time measured Analyte concentration Method details ...

Page 152: ......

Page 153: ...hlength adjustment and a single point baseline correction can also be used To make UV Vis measurements Measures the absorbance of any sample at up to 40 wavelengths across the ultra violet UV and visible regions of the spectrum Measure UV Vis Reported Results Settings Detection Limits NOTICE Do not use a squirt or spray bottle on or near the instrument as liquids will flow into the instrument and ...

Page 154: ... holder Tip If using a cuvette make sure to align the cuvette light path with the instrument light path 4 Tap Blank and wait for the measurement to complete Tip If Auto Blank is On the blank measurement starts automatically after you lower the arm This option is not available for cuvette measurements 5 Lift the arm and clean both pedestals with a new laboratory wipe or remove the blanking cuvette ...

Page 155: ...r application See Choosing and Measuring a Blank for more information For micro volume measurements Ensure pedestal surfaces are properly cleaned and conditioned Ensure samples are homogeneous before taking a measurement Avoid introducing bubbles when mixing and pipetting Follow best practices for micro volume measurements Use a 1 2 μL sample volume See Recommended Sample Volumes for more informat...

Page 156: ... measurement details Drag tab down up to see more less sample data Swipe screen left to view table with more measurement results Pinch and zoom to adjust axes double tap to reset Tap to end experiment and export data UV Vis spectrum Tap to edit Menu of options tap to open Sample name tap to edit Absorbance at user defined wavelength 1 450 nm Absorbance at user defined wavelength 2 623 nm Tap to ed...

Page 157: ...ress and hold the sample row Here is an example Related Topics Basic Instrument Operations Note Scroll up to display absorbance values for any additional user defined wavelengths Sample name tap to edit Sampling method Application Date time measured Automated pathlength setting Baseline correction absorbance Absorbance at 450 nm Baseline correction wavelength Absorbance at 623 nm Absorbance at 635...

Page 158: ...ed pathlength selection Allows the software to use the optimal shorter pedestal pathlength for high concentration samples to help prevent detector saturation see Detection Limits for details When selected the shorter pathlength is used when any wavelength between 220 nm and 850 nm has 10 mm equivalent absorbance value of 12 5 or higher For wavelengths between 190 nm and 219 nm the change to the sh...

Page 159: ...Measure UV Vis Thermo Scientific NanoDrop One User Guide 153 ...

Page 160: ...Measure UV Vis 154 NanoDrop One User Guide Thermo Scientific ...

Page 161: ...37 C heater and micro stirrer To make kinetic measurements Make time based kinetic measurements using the cuvette holder NanoDrop OneC model instruments only Measure Kinetics Create Kinetics Method Edit Kinetics Method Reported Results Settings Detection Limits Note The instrument arm can be up during cuvette measurements which allows you to add reagents to the sample solution if desired NOTICE To...

Page 162: ...he method name in the Select Method box create a new method by tapping Create Method specifying the method settings and choosing Save Method edit an existing method by tapping the method name and choosing Edit Method 3 Specify any cuvette options such as heating or stirring by tapping Settings see General settings for details Note If your cuvette pathlength is not 10 mm specify the correct pathlen...

Page 163: ...s you the current temperature and waits for the heater to reach the target temperature before starting the measurements Note You may add reagents to the sample solution at any time during the measurement Use the Pause button at the bottom of the measurement screen to pause the experiment if you need to end the experiment early tap Stop Wait for all measurement stages to complete Remove cuvette and...

Page 164: ...rument However once the method is created it can be saved in the NanoDrop One database on the local instrument or in the NanoDrop Viewer database on a connected PC To create a new kinetics method from Home screen tap Kinetics tab Kinetics application tap Create Method the method settings are displayed with Name and Range tab selected enter Method Name and Description if desired select Measurement ...

Page 165: ...age the colored boxes correspond with the specified delay and number of intervals for each stage to save the method and return to the Kinetics menu tap Save Method to run the method tap Run Method Related Topics Edit Kinetics Method Edit Kinetics Method Kinetics methods can be edited only on the NanoDrop One instrument To edit an existing kinetics method if the instrument has a connected PC Ethern...

Page 166: ...screen tap Kinetics tab Kinetics application select a method by tapping the method name in the Select Method box tap Edit Method edit method settings as desired tap Save Method to save your changes tap Run Method to run the updated method Related Topics Create Kinetic Method ...

Page 167: ...down to see more entries Each item in the list at the right has a corresponding absorbance spectrum at the left The image below highlights the available features Tap row to select measurement and update spectrum tap more rows to overlay up to five spectra Press and hold row to view measurement details Drag tab down up to see more less sample data Swipe screen left to view Rate measurement screen P...

Page 168: ...ime with time on the X axis and absorbance on the Y axis Measurements taken at each specified wavelength are presented in a unique color A key showing the monitored wavelengths and their assigned colors appears in the upper left corner of the screen Tap to update plot to show change in measured absorbance over time see example below Swipe screen left to view data table swipe right to return to abs...

Page 169: ...sorbance over time where each data point is the difference in absorbance from the previous measurement Tap to update plot to show measured absorbance over time see example above Swipe screen left to view data table swipe right to return to absorbance measurement screen Tap to end experiment and export data User defined wavelengths ...

Page 170: ... defined wavelengths at a given stage and time Scroll down to see measurement information that is out of view The image below highlights the available features Swipe screen right to return to Rate measurement screen Tap to end experiment and export data Absorbance values for each user defined wavelength Press and hold row to view measurement details Measurement number Stage Measurement time click ...

Page 171: ... and hold the measurement row Here is an example Related Topics Basic Instrument Operations Measurement number Sampling method Date time measured Meas stage Absorbance at 260 nm Absorbance at 340 nm Absorbance at 660 nm User defined wavelengths Meas time Method details scroll up to view more Application used Delete this measurement Return to previous screen Printthis screen ...

Page 172: ...hod name Enter a name for this method this name appears in the Kinetics Setup box after the method has been saved Description Enter a detailed description for this method if desired such as the type of samples added reagents etc Measurement range Select the spectral range in which this method will acquire data Available options Ultra violet only 190 nm 350 nm Visible only 350 nm 850 nm Ultra viole...

Page 173: ...of absorbance measurements to take in this stage Note Since the first measurement is taken when the stage starts the number of measurements reported for each stage will be the Intervals setting plus 1 Duration Readout shows the total time required for this stage including any delay and all specified intervals The colored rows at the right see image below show the start and end times for each stage...

Page 174: ...ds Stage 2 9 11 13 15 and 17 seconds Stage 3 22 27 and 32 seconds Note Kinetic experiments are limited to 1000 measurements This means the total number of measurements from all intervals in all stages must be less than 1000 Consider available instrument or computer disc space for lengthy experiments Tab Setting Description ...

Page 175: ...Learning Center How the Instrument Works Set Up the Instrument Measure a Micro Volume Sample Measure using a Cuvette Prepare Samples and Blanks Basic Instrument Operations Acclaro Sample Intelligence Instrument Settings NanoDrop One Viewer Multimedia ...

Page 176: ...ter uses surface tension to hold a small volume of sample between two pedestals The patented sample retention system enables the measurement of highly concentrated samples without the need for dilutions A fiber optic cable embedded in the upper pedestal leads to a xenon light source A second cable embedded in the lower pedestal leads to a detector When the instrument arm is down the sample forms a...

Page 177: ...lculate the total absorbance of the sample according to the equation at the left Beer Lambert equation where A absorbance in absorbance units A wavelength dependent molar absorptivity coefficient or extinction coefficient in liter mol cm b pathlength in cm c analyte concentration in moles liter or molarity M Sample Concentration The Beer Lambert equation Beer s law shown at the left is used to cor...

Page 178: ...an Accessory To connect a compatible printer or other compatible accessory such as a USB keyboard and or mouse to the instrument use any USB port on the instrument front back left or back right See Accessories for information about accessories compatible with the NanoDrop One instruments Ethernet Power on off Power USB A 2 CAUTION Avoid shock hazard Each wall outlet used must be equipped with a gr...

Page 179: ...d no Bluetooth input devices are listed tap Off button to enable Bluetooth connectivity button turns blue changes to On and software automatically searches for any available Bluetooth input devices If no Bluetooth devices are found after a few seconds the message No nearby Bluetooth devices were found is displayed to add a Bluetooth device follow manufacturer instructions to pair the device for ex...

Page 180: ...e Devices list a pairing request similar to the following may be displayed complete any instructions to pair the device Note If your Bluetooth device does not pair restart the device and then repeat the steps above to pair it with the instrument you may also try turning Bluetooth off and back on After a device is paired it remains paired even after the instrument is restarted ...

Page 181: ... Bluetooth device for input without disconnecting or unpairing it This allows others to easily reselect and use the device for input For example if there are multiple connected and paired Bluetooth input devices such as a keyboard and a barcode scanner follow these steps to select the devices to use or to deselect devices you don t want to use from instrument Home screen tap tap System tab tap Blu...

Page 182: ...k to return to System settings tap Done to close Settings Disconnect Bluetooth device from instrument Home screen tap tap System tab tap Bluetooth to disconnect paired Bluetooth device tap its Profiles button Note If no Bluetooth device is selected for input the instrument relies on the integrated touchscreen keyboard for input To select the device again follow the steps above and select the devic...

Page 183: ...ter can then be used to store or view data acquired with the NanoDrop One instrument NanoDrop One Viewer software must be installed on the computer Tools needed Standard straight through Ethernet cable CAT5e or newer is recommended Set up Ethernet connection connect Ethernet cable between Ethernet port on instrument back panel see image above and Ethernet port on computer Note If the computer is a...

Page 184: ...een tap Settings tap System tab tap Wi Fi if Wi Fi is disabled button in upper right is set to OFF and no wireless networks are listed tap button to enable Wi Fi and display available Wi Fi networks select remote computer s Wi Fi network host and tap Connect here is an example Note To store or view collected data on a connected computer using W Fi NanoDrop One Viewer software must be installed on ...

Page 185: ... at the right of the Wi Fi button as in the example below record IP address you will need to enter it on the remote computer in the next section tap Done to exit Settings Note Some Wi Fi networks may require an identity password or other information before you can connect to them or they may be anonymous that is you may have to search for them by name For more information see the system administra...

Page 186: ...played enter the following information Instrument IP address displayed on instrument in Settings System see previous section if IP address is valid Status column shows Valid IP Address unique Instrument Name in case there are multiple instruments in the same lab on the same network PC Name such as the computer s assigned name or an invented name the name you enter will appear in the select a data ...

Page 187: ...m the list tap to select the row and then tap Delete when finished choose OK to close Wi Fi Data Storage setup NOTICE You can add multiple Wi Fi addresses to this list to make it easy to switch computers However only one Wi Fi connection should be enabled during data collection to ensure the data integrity Instrument information Add Wi Fi connection with a new instrument Delete selected Wi Fi conn...

Page 188: ...to the instrument with an Ethernet cable set Data Storage to Direct Connect PC see Set Up Ethernet Connection for details to store all subsequently acquired measurement results in the NanoDrop One Viewer database on a computer connected to the instrument through a wireless network set Data Storage to the computer s assigned name see Set Up Wi Fi Connections for details The Ethernet and wireless op...

Page 189: ...onnection had already been established to close message box new data storage location appears adjacent to Connectivity Status icon all subsequently acquired measurement results are saved in the NanoDrop Viewer database on the selected computer and in the NanoDrop One database on the local instrument PC name Personal computer PC IP address Data storage location wireless computer Computer sIP addres...

Page 190: ...ment to be saved there If the wireless or Ethernet connection is interrupted during a measurement data storage switches back to the local instrument with no loss of data Custom methods and Kinetics methods must reside on the connected computer when the data are configured for remote storage When the instrument is connected to a computer with an Ethernet cable or through a wireless network the Data...

Page 191: ... 35 C 41 F 95 F relative humidity non condensing 20 80 Locate the instrument away from air vents and exhaust fans to minimize evaporation After the instrument is installed you can leave it turned on Related Topics Safety and Operating Precautions Instrument Models and Features Location of database where instrument is currently storing data Local instrument or Connected PC Wi Fi status Bluetooth st...

Page 192: ...3 Learning Center Set Up the Instrument 192 NanoDrop One User Guide Thermo Scientific Optional Accessories Instrument Settings ...

Page 193: ...retention system enables the measurement of highly concentrated samples without the need for dilutions Tap here for details Supplies needed NanoDrop One or NanoDrop OneC spectrophotometer lint free laboratory wipes calibrated precision pipettor 0 2 μL sample material resuspended in appropriate buffer solution see Preparing Samples pure buffer solution for blanking instrument see Choosing and Measu...

Page 194: ...tion Before first measurement clean both pedestals with a new laboratory wipe Run a blanking cycle to verify pedestals are clean After each measurement clean both pedestals with new wipe to prevent carryover After each set of measurements clean pedestals with DI H2O see Clean pedestals between users Recondition pedestals periodically to maintain their hydrophobic property ...

Page 195: ... measurement If using low accuracy 0 10 μL pipettor use 2 μL sample volumes Use new tip for each blank and sample aliquot Use new aliquot of sample for each measurement If solvents are used make sure they are compatible with the pedestals see Compatible Solvents in Hazardous Materials Application Sample Volume Nucleic acid aqueous solution 1 μLa a Use 2 μL for samples that contain materials that m...

Page 196: ...z fiber optic cables 1 If the instrument has a working Ethernet or wireless connection to a personal computer PC the Connectivity Status icon is blue and shows the currently selected location for storing and viewing data collected with the instrument If the Connectivity Status icon is blue tap the icon and set Data Storage to Local as shown at the left 2 From the instrument Home screen select an a...

Page 197: ...ement to complete Tip If Auto Blank is On blank measurement starts automatically after you lower the arm Lift the arm and clean both pedestals with a new laboratory wipe 5 Measure the first sample Pipette 1 2 μL sample solution onto the pedestal and quickly lower the arm see Recommended Sample Volumes for more information Start the sample measurement if Auto Measure is On lower arm if Auto Measure...

Page 198: ...vailable on demand technical support available invalid result 6 To measure another sample Lift the arm Clean both pedestals with new wipe Load the next sample and quickly lower the arm Start the sample measurement Wait for the measurement to complete The new spectrum replaces the previous one on the spectral display and the new reported values appear under the previous ones in the table Drag tab d...

Page 199: ...e box use displayed keyboard to type name tap Done key or leave the default experiment name Tap Lift the arm and clean both pedestals with a new wipe If finished with the instrument for the day clean the pedestals with DI H2O see Clean pedestals between users Acquired data are automatically saved in an experiment with the entered name In the default configuration experiments are stored in a databa...

Page 200: ...ell cultures and kinetic studies The cuvette system offers an extended lower detection limit and an optional 37 C heater and micro stirrer Supplies needed NanoDrop OneC spectrophotometer lint free laboratory wipes two compatible cuvettes sample material resuspended in appropriate buffer solution see Preparing Samples pure buffer solution for blanking instrument see Choosing and Measuring a Blank o...

Page 201: ...re samples with analysis wavelengths in the UV range 340 nm Micro semi micro and ultra micro cuvettes should be masked Fill cuvettes with enough blanking or sample solution to cover instrument optical path 2 mm sample beam is 8 5 mm above cuvette bottom Lift instrument arm and make sure cuvette holder is free of debris When inserting quartz or masked plastic cuvettes align cuvette light path with ...

Page 202: ...tte set Pathlength to pathlength width of cuvette see cuvette manufacturer for specifications set stirrer and heater if desired tap Done See General settings for details 3 Measure a blank Fill clean dry cuvette with enough blanking solution to cover instrument optical path Lift instrument arm and insert blanking cuvette into cuvette holder making sure to align light path of cuvette with light path...

Page 203: ... Blanks Acclaro Sample Intelligence Instrument Settings Search Experiment Database Export Data Measure a Micro Volume Sample 4 Measure a sample Fill clean cuvette to same height with sample solution Replace blanking cuvette with sample cuvette making sure to align light paths Tap Measure Wait for measurement to complete Remove cuvette Clean cuvette according to manufacturer specifications ...

Page 204: ...e protocol After purification analyte of interest is typically dissolved in aqueous buffer solution before it is measured Tip Any molecule that absorbs light at analysis wavelength will contribute to total absorbance value used to calculate sample concentration Ensure final analyte concentration is within instrument s absorbance detection limits For micro volume measurements gently but thoroughly ...

Page 205: ...specially at the analysis wavelength as in the example at the right If the resulting spectrum is greater than 0 04 A around the analysis wavelength that buffer solution may interfere with the sample analyses especially for low concentration samples See below for details Good blanking buffer measured abs 0 04 Problems associated with blanking Residual sample was left on pedestal or in cuvette befor...

Page 206: ...tch multimedia training Evaluating a Blanking Solution for Suitability Solutions for blanking problems Thoroughly clean and or recondition both pedestals and then rerun blanking cycle or measure new blank using new aliquot of appropriate buffer solution then measure new aliquot of unknown sample For most applications blank with the same buffer solution used to resuspend the analyte of interest The...

Page 207: ...ne User Guide 207 NOTICE Do not use a squirt or spray bottle on or near the instrument as liquids will flow into the instrument and may cause permanent damage Do not use hydrofluoric acid HF on the pedestals Fluoride ions will permanently damage the quartz fiber optic cables ...

Page 208: ...fer solution Pipette 1 2 μL buffer solution onto the pedestal and lower the arm Start the sample measurement if Auto Measure is On lower arm if Auto Measure is off lower arm and tap Measure Wait for measurement to complete The resulting spectrum should vary no more than 0 04 A from the baseline at the analysis wavelength 260 nm for nucleic acids 280 nm for proteins If your spectrum does not meet t...

Page 209: ...nd Blanks Thermo Scientific NanoDrop One User Guide 209 Effects of Bubbles in Samples What is a Blank Evaluating a Blanking Solution for Suitability Maintaining the Pedestals Measure a Micro Volume Sample Measure a Sample Using a Cuvette ...

Page 210: ...3 Learning Center Basic Instrument Operations 210 NanoDrop One User Guide Thermo Scientific Basic Instrument Operations Home Screen Measurement screens Data Viewer General Operations ...

Page 211: ...ications The NanoDrop One offers a broad range of applications for measuring samples with the instrument To select an application tap an Application tab such as Nucleic Acids and then tap an application name such as dsDNA Tap here for detailed information about each available application Data Viewer Instrument Settings Instrument Diagnostics System Status Applications Training and Help ...

Page 212: ... where instrument is currently storing data These options are available Local instrument Connected PC personal computer connected through Ethernet cable or wireless network the Ethernet and wireless options listed above also store data on the instrument as a backup Wi Fi status Status of WiFi connections for the instrument Connected to Enabled and not connected or Disabled Bluetooth status Status ...

Page 213: ...tings Instrument Diagnostics Tap on the Home screen to verify instrument operation Instrument diagnostics should be run periodically according to the recommended maintenance schedule Tap here for information about how to run the available instrument diagnostics Firmware version Version of instrument firmware installed Android release Custom version of Android release Android version Version of And...

Page 214: ...tem The NanoDrop One software comes with comprehensive embedded training and support Tap here for information on how to navigate the available information Related Topics Applications Set Up the Instrument NanoDrop One Data Viewer Instrument Settings Instrument Diagnostics About this Help System Measure a Micro Volume Sample Measure a Sample Using a Cuvette ...

Page 215: ...less sample data Pinch and zoom to adjust axes Tap to end experiment and export data Sample name tap to edit UV absorbance spectrum for selected sample Menu of options tap to open Measurement alert tap to learn more Tap to measure blank solution Page control swipe screen left to view table with more measurement results Tap to measure sample solution Selected application Auto Measure Measurement re...

Page 216: ...elow In this example the first sample would be named Sample 1 followed by Sample 2 etc You can edit the default base name and overwrite any sample name Edit default sample base name After you measure a blank and before the first sample is measured tap Sample Name field to display keyboard enter new base name tap Done key Edit sample name from Home screen tap to open Data Viewer select experiment s...

Page 217: ...ported Results Here is an example for dsDNA Absorbance Spectrum For each measured sample each application shows the UV or UV visible absorbance spectrum and a summary of the results The vertical axis shows absorbance in absorbance units A The horizontal axis shows wavelength in nm Sample Pathlength All applications display the sample pathlength along the spectrum s vertical axis Micro volume absor...

Page 218: ...d to resuspend the sample For more information see Choosing and Measuring a Blank Measure Tap Measure to measure a sample for the selected experiment Samples must be properly isolated and prepared before they can be measured with the instrument and the concentration must be within the instrument s absorbance detection limits For more information see Preparing Samples and Measure a Micro Volume Sam...

Page 219: ...ect or deselect Auto Measure from any sample measurement screen tap the On or Off button at the right of the Measure button Auto Blank To select or deselect Auto Blank from any blank measurement screen tap the On or Off button at the right of the Blank button End Experiment Tap End Experiment when you are ready to name and save your experiment add a label to help you locate the experiment later se...

Page 220: ...c Sample Details Press and hold a sample row in any measurement screen or data table to show the sample details which include all available measurement results and associated details for the selected sample Here is an example Note You can also edit the sample name from the Sample Details box ...

Page 221: ...the experiment The image below highlights the available features Related Topics Measure Nucleic Acids Measure Proteins Instrument Settings Print Data Acclaro Sample Intelligence Tap row to select sample Press and hold row for sample details Measurement alert tap to learn more Application used Page control swipe screen right to return to measurement screen Menu of options tap to open Sample name ta...

Page 222: ...ccording to acquisition date experiment name application used and any assigned labels see Manage Identifiers From the Data Viewer you can locate and select any experiment to see the measurement data it contains or export selected experiments to a variety of locations and formats These operations are available from the Data Viewer Search for an experiment or change time range filter Tap row to view...

Page 223: ...strument in order to view acquired spectra and associated data from any experiment at any time Open instrument database of measurement results to open NanoDrop One database on instrument tap Data Viewer on instrument Home screen Menu Tap in the Data Viewer to see the available menu options Note The Data Viewer icon is not available on the instrument Home screen when the instrument is connected to ...

Page 224: ...e Identifiers for information about adding and deleting labels Here is an example Export Selected Experiments Use Select in the Data Viewer to select experiments to be exported Export selected experiments tap row in Data Viewer to list experiments acquired on that date or use Search feature to find experiment insert USB memory device into available USB port on instrument front back left or back ri...

Page 225: ...perations for details tap Export after Export Success message tap OK Delete Selected Experiments Use Select in the Data Viewer to select experiments to be deleted Delete selected experiments tap row in Data Viewer to list experiments acquired on that date or use Search feature to find desired experiment tap Select tap to select one or more experiments to delete tap again to deselect an experiment ...

Page 226: ...ature to find desired experiment tap experiment name to open the experiment Here is an example The Data Viewer provides measurement data as spectral data and data tables similar to what you see after you complete a measurement Note The data shown are dependent upon the application used to measure the samples nucleic acids in these examples For more information see the application details Nine expe...

Page 227: ...age below describes the available features Drag tab down up to see more less sample data Page control swipe screen left or right to view next or previous screen Pinch and zoom to adjust axes Tap row to select sample and update spectrum tap more rows to overlay up to five spectra Press and hold sample row to view sample details Selected experiment Measurement alert tap to learn more Selected applic...

Page 228: ...ct sample Press and hold row for sample details Selected experiment Measurement alert tap to learn more Application used Page control swipe screen right to view previous screens 2 Menu of options tap to open Measurement results see Applications for details Tap to select unit Home Return to NanoDrop One Home screen Manage Identifiers Add or delete labels for selected experiment to make it easier to...

Page 229: ...ing on the instrument or from the NanoDrop One Viewer software installed on a personal computer see Manage Identifiers on a PC Use the Data Viewer to add labels to experiments assign existing labels view assigned labels and remove or delete labels on the instrument You can filter the list of experiments in the Data Viewer based on one or more user defined labels Label new experiment when you save ...

Page 230: ...ss and hold selected experiment to see Experiment Details Find labeled experiments from Home screen tap to open Data Viewer tap Search in Search box select date range select application only applications that have associated data are shown select one or more identifiers from scrollable list and tap OK Remove a label from Home screen tap to open Data Viewer tap to open an experiment tap and choose ...

Page 231: ... all the selected experiments Use any spreadsheet or word processing application to open a CSV or TSV file Here is an example of several sample measurement results in CSV format The SQL file can be opened only using our NanoDrop One Viewer software and only after the file has been imported Data can be exported to a USB device connected to any USB port on the local instrument front back left or bac...

Page 232: ...port Success message tap OK remove USB device tap End Experiment Export data from Data Viewer from Home screen tap to open Data Viewer tap row in Data Viewer to list experiments acquired on that date or use Search feature to find experiment insert USB memory device into available USB port on instrument front back left or back right tap Select tap to select one or more experiments to export tap aga...

Page 233: ...ou can delete a sample measurement from any experiment Delete data from any measurement screen press and hold sample row to open Sample Details box tap Delete data from Data Viewer from Home screen tap to open Data Viewer tap row in Data Viewer to list experiments acquired on that date or use Search feature to find desired experiment press and hold sample row to open Sample Details box tap NOTICE ...

Page 234: ...eens tap to select one or more sample rows to print tap again to deselect a sample row tap and choose Print in the Print Information box choose OK One label is printed for each selected measurement Print data from Data Viewer from Home screen tap to open Data Viewer tap row in Data Viewer to list experiments acquired on that date or use Search feature to find desired experiment tap experiment name...

Page 235: ...data table in any measurement screen or from the Data Viewer press and hold sample row to open Sample Details box tap in the Print Information box choose OK One label is printed for this measurement Related Topics Instrument Settings NanoDrop One Data Viewer Search Experiment Database Select Experiments to Export or Delete Open Experiment ...

Page 236: ...hoose Settings These instrument settings are available Wi Fi Set up wireless local area network WLAN connection on the instrument Bluetooth Set up Bluetooth connections to wireless input devices for the instrument such as a wireless keyboard mouse or barcode scanner Language Select language for displaying NanoDrop One software and for any connected input device such as a keyboard mouse or barcode ...

Page 237: ... Time Zone is selected Use 24 hour format use 24 hour time format Choose date format choose an available date format Brightness Adjust brightness of instrument touchscreen Sound Volume Adjust volume of instrument touchscreen Update Software Update NanoDrop One software via USB device connected to instrument if connected USB device contains multiple eligible update files you can choose which files ...

Page 238: ... Up Ethernet Connection Dye Chromophore Editor Protein Editor Use cuvette Select cuvette sampling mode available for NanoDrop OneC instrument model only When selected these additional options are available Pathlength Enter cuvette pathlength width before taking blank or sample measurements with cuvettes see cuvette manufacturer for cuvette specifications Stir Speed If using automatic stirring drop...

Page 239: ...3 Learning Center Instrument Settings Thermo Scientific NanoDrop One User Guide 239 ...

Page 240: ...as a resource for further study and a learning tool for new or novice users The Thermo Scientific Acclaro Sample Intelligence technology built into the NanoDrop One instruments provides these exclusive features to help you assess sample integrity contaminant analysis to help qualify a sample before use in downstream applications on demand technical support for measurements that are atypical or ver...

Page 241: ...ally see examples below Tap the icon to review the associated data or information The icons appear next to the measurement results see above and in the data table as well as the Data Viewer see below The icons are active in all three places the information remains with the data indefinitely even after it has been exported Contaminant analysis is available for this measurement Technical information...

Page 242: ...ing the measurement Examples of known contaminants include for dsDNA and RNA measurements in the analysis region protein and phenol also detects presence of guanidine HCl and guanidinium isothiocyanate for protein measurements in the analysis region nucleic acids and phenol If contaminants are identified in a sample the Contaminant Analysis icon appears to the left of the measurement results Tap t...

Page 243: ...A260 A280 and A260 A230 ratios out of range and caused the reported nucleic acid concentration to be higher than the real value The software identifies the impurity protein and reports the following baseline corrected absorbance due to protein 2 83 at the analysis wavelength 260 nm coefficient of variation for the measurement result uncertainty x 100 measurement result 5 3 a high CV indicates the ...

Page 244: ...ination When a purity ratio falls outside the expected range the spectral profile is often examined qualitatively Our Acclaro technology applies a quantitative approach to contaminant analysis Through sophisticated mathematical algorithms Acclaro analyzes the spectral data to identify probable contaminants in a sample and removes any contribution due to the contaminant from the sample result This ...

Page 245: ...h sample measurements also referred to as purity ratios For more information watch the multimedia training What is a Purity Ratio bubble check which looks for bubbles or other reflective materials in a sample or blank For more information watch the multimedia training Effects of Bubbles in Samples If technical information is available the information icon appears to the left of the measurement res...

Page 246: ...tion provided for the bubble error Tap Learn More for the next level of information which for this example includes a link to a multimedia training video Invalid Results Alerts The NanoDrop One software uses an embedded image sensor to monitor all measurements for conditions such as a broken liquid column that are likely to invalidate the measurement results ...

Page 247: ...uide 247 After an invalid results alert the Invalid Results icon is displayed and the measurement is stopped See Troubleshooting for more information Related Topics NanoDrop One Measurement Screens NanoDrop One Data Viewer Maintenance Schedule Maintaining the Pedestals Troubleshooting ...

Page 248: ...ata from the Instrument below or saved directly on a connected computer after each measurement is completed see Set Up Ethernet Connections or Set Up Wi Fi Connections in Set Up the Instrument for details Install the Viewer software The Viewer software can be installed on any computer running the Windows operating system software See our website for compatible versions of Windows software To downl...

Page 249: ...hods menu option File menu Help menu Time range search filter Data search filters Refresh Custom Methods menu Import Data For importing experiments from the instrument Set Up Wi Fi Data Storage Set up this computer as a potential data storage location for samples measured with the instrument see Set Up Wi Fi Connections in Set Up the Instrument for more information Exit Closes the Viewer software ...

Page 250: ...tion date for example last six months or in a specific date range To turn off the time range filter set it to All For more information see Search Viewer Database Refresh Updates the list of experiments and measurement results in the Viewer software after new data have been imported Right Click Menu Right click an experiment for these additional menu options NanoDrop One Website Opens a local web b...

Page 251: ...l computer that has NanoDrop One Viewer software installed from Viewer software choose File Import Data navigate to portable USB device select exported SQL file and choose Open Export Experiment Export spectra and results for selected measurements in several formats Manage Identifiers Add user defined labels to experiments view assigned labels and remove or delete labels Experiment Details Display...

Page 252: ...earch experiment list To search the experiment list change a filter setting on the Viewer Home screen The list is updated automatically These filters are available Application type Only applications that have associated experiments in the Viewer database are available To list only the experiments acquired using the OD600 application for example select OD600 under the Application Type filter select...

Page 253: ...tifier filter make sure none of the buttons are selected Time range To list only the experiments acquired during a particular date range during the last six months for example select the date range from the drop down list To turn off the time range filter set it to All Here is an example of database filtering Click to select or deselect application filters Change a filter to display updated list o...

Page 254: ...database in order to view print or export the spectra and associated data The Viewer Home screen shows a list of experiments that match the current filter settings Open experiment use available filters to locate the experiment see Search Viewer Database for details double click the experiment name in the filtered experiment list the experiment opens and the software displays the measurement screen...

Page 255: ...screen Export Experiment details Manage Identifiers Print Spectral pane Click to change displayed unit Spectrum Click and hold a point to show absorbance value Right click to overlay spectra Measurement details see individual Applications for more information Use mouse scroll wheel to expand or contract spectrum double click to reset Click to edit sample name Experiment name ...

Page 256: ...asurements use Ctl click to select nonsequential measurements click The Print Report window is displayed with a preview of the first page along with these options These standard Windows Print options are also available Page Setup to select paper size and adjust margins and page orientation Print Preview allows all pages to be previewed click Print select a printer click Print Print spectra Print t...

Page 257: ...e can be opened only using our NanoDrop One Viewer software and only after the file has been imported The XML file can also be opened with an XML reader application Export selected measurements open the experiment click to select a measurement to export use Shift click to select sequential measurements use Ctl click to select nonsequential measurements click in Export Experiments box navigate to a...

Page 258: ...orbance value are displayed in a popup box Overlay Spectra If you want to display the spectra for all measurements in the experiment layered on top of each other in the spectral pane open the experiment right click the spectral pane and select Overlay Mode The spectra are displayed in a variety of colors NOTICE Deleted data cannot be recovered Note You can also drag the mouse along the spectrum s ...

Page 259: ...plications offer a choice of concentration units Change concentration unit for selected experiment open the experiment use the drop down menu if available in the measurement table to change the displayed unit After the unit is changed all reported concentration results are recalculated to comply with the new unit View Experiment Details You can display information about the open experiment includi...

Page 260: ...the instrument see Manage identifiers on the instrument or from the NanoDrop One Viewer software installed on a personal computer Use the Viewer software to add labels to experiments assign existing labels view assigned labels and remove or delete labels on a personal computer You can filter the list of experiments based on one or more user defined labels Show or Hide User Identifier Filter Button...

Page 261: ...rrow points down Label Experiment in Viewer You can use the Viewer software to add user defined labels to experiments The list of experiments can then be filtered based on those labels see Find labeled experiment for details Add new label to experiment from Viewer Home screen right click experiment in experiment list and choose Manage Identifiers in Manage Identifiers box enter label and tap or pr...

Page 262: ...r Home screen Assign existing label to experiment from Viewer Home screen right click experiment in experiment list and choose Manage Identifiers in Manage Identifiers box start typing existing label name actual name of existing label appears below entry box select auto filled existing label and choose OK existing label is added to selected experiment New label ...

Page 263: ...etails choose OK Find Labeled Experiments You can use the Viewer software to find experiments with assigned user defined labels The database is automatically filtered using the current filter settings Find experiments that have a particular user defined label from Viewer Home screen set search filters deselect any Application Type filters if desired so the experiments acquired with all application...

Page 264: ... the Viewer software to easily remove a user defined label from an experiment A removed label remains in the User Identifier Filter list if it is still assigned to other experiments Remove a label from Viewer Home screen right click experiment in experiment list and choose Manage Identifiers in Manage Identifiers box select one or more label buttons selected label buttons are blue tap selected lab...

Page 265: ...data with the instrument Custom methods can be made with or without standards Create Custom Method Create method to be used for sample measurements with user defined settings Create new custom method from Viewer Home screen choose Custom Methods menu Manage Custom Methods in Manage Custom Methods box choose one of the following Create Formula Method if your method will not have standards Create St...

Page 266: ... Setup box on the instrument after the method has been transferred there enter detailed Description of method if desired specify how to calculate and report the method results if method does not have standards specify factor or extinction coefficient of analyte enter 1 to report absorbance measurements only Custom Method Setup for Create Formula Method selection ...

Page 267: ...ect the curve fit type enter or choose remaining custom settings as needed see below choose Save if the method has a green check mark icon at the top tap Close to exit method setup Note If this icon appears next to the Save button instead of a green check mark icon the method is invalid because it contains an error Hover your mouse over the icon for suggested solutions ...

Page 268: ...68 NanoDrop One User Guide Thermo Scientific View or edit custom method choose Custom Methods menu Manage Custom Methods existing methods are listed in Select Method box along with their type formula or standards and Description custom formula method ...

Page 269: ...n and or analysis correction wavelength For micro volume absorbance measurements and measurements taken with nonstandard other than 10 mm cuvettes the spectra are normalized to a 10 mm pathlength equivalent Analysis wavelength Monitor absorbance at specified wavelength enter the wavelength in nanometers Note The specified wavelength must fall within the selected measurement range The measurement r...

Page 270: ...ctor typically 1 where wavelength dependent molar absorptivity coefficient or extinction coefficient b pathlength in cm determined at measurement time then normalized to 10 mm 1 cm pathlength equivalent Extinction coefficient and molecular weight Enter extinction coefficient for 1 cm pathlength and use adjacent drop down list to select appropriate unit Equation below shows how extinction coefficie...

Page 271: ...urve can be generated using two or more standards The software allows a reference and up to 7 standards All reference and standards solutions should be in the same buffer used to resuspend the samples plus the same volume of reagent added to the samples First standard can be a reference measurement The reference solution should contain none of the analyte of interest The reference measurement is n...

Page 272: ...ght scattering particulates by subtracting the absorbance at a specified baseline point Then specify wavelength for baseline correction Note Software subtracts absorbance value at specified baseline correction wavelength from absorbance values at all wavelengths in sample spectrum As a result absorbance of sample spectrum is zero at specified baseline correction wavelength Analysis wavelength corr...

Page 273: ... used Recommended for samples that are highly absorbing at the analysis wavelength This option may cause reduced sensitivity when the sample spectra have a large absorbance peak that is not at the analysis wavelength Note When the analysis wavelength is between 190 nm and 219 nm the optimal longer pathlength is used when sample absorbance is less than or equal to 10 10 mm pathlength equivalent and...

Page 274: ...is reported in Data Table and Sample Details screens Unit Enter unit for reported result After a measurement the unit is reported in Data Table and Sample Details screens Edit Edit selected formula for current method Delete Delete selected formula from current method Formula rules Custom formulas can include the following operators and functions Path Returns sample pathlength in cm A nm Returns sa...

Page 275: ...d if your instrument is not connected to the computer with an Ethernet cable or through a wireless network Export Custom Method Export a custom method in order to run it and store the measurement results on the NanoDrop One instrument from Manage Custom Methods box select custom method Note If the computer is connected to the instrument with an Ethernet cable or through a wireless network custom m...

Page 276: ...USB memory device and then load the method see To Load a Custom Method for details Import custom method Import a custom method back to a computer running the NanoDrop One Viewer software in order to edit the method settings from Manage Custom Methods box choose Import locate and select method file choose Open imported method is added to end of Select Method list Edit custom method Edit a custom me...

Page 277: ...3 Learning Center Multimedia Thermo Scientific NanoDrop One User Guide 277 Multimedia What is a Purity Ratio Evaluating a Blanking Solution for Suitability What is a Blank Effects of Bubbles in Samples ...

Page 278: ...3 Learning Center Multimedia 278 NanoDrop One User Guide Thermo Scientific Pedestal Cleaning and Reconditioning ...

Page 279: ...Thermo Scientific NanoDrop One User Guide 279 4 Maintaining Your Instrument Maintenance Schedule Clean Touchscreen Maintain Pedestals Decontaminate Cuvette System Instrument Diagnostics ...

Page 280: ...h 0 5M HCl Recondition pedestals Every 6 Months Recondition pedestals Run Intensity Check Run Performance Verification Run Pedestal Image Check If you are experiencing an issue with your system refer to the troubleshooting information If the issue persists contact us If you are outside the U S A and Canada please contact your local distributor If your instrument requires maintenance or repair cont...

Page 281: ...material such as paper towel apply excessive pressure spray liquid directly onto the touchscreen apply lubricant to the touchscreen slide mechanism To clean the touchscreen Gently wipe the touchscreen with a soft lint free cloth such as microfiber If necessary use a cleaner intended for glass LCD displays and follow the manufacturer s recommendations Related Topics Clean Pedestals Recondition Pede...

Page 282: ...may be required for periodic maintenance Clean Pedestals Recondition Pedestals NOTICE Do not use hydrofluoric acid HF on the pedestals Fluoride ions will permanently damage the quartz fiber optic cables To prevent damage from spills keep containers of liquids away from the instrument Do not use a squirt or spray bottle on or near the instrument as liquids will flow into the instrument and may caus...

Page 283: ...ntaining the Pedestals Thermo Scientific NanoDrop One User Guide 283 Note Solutions containing detergent or isopropyl alcohol may uncondition the pedestals If these are required for sample analyses follow immediately with 3 5 μL DI H2O ...

Page 284: ...al with a new laboratory wipe To clean the pedestals between users 1 Lift the arm and clean both pedestals with a new laboratory wipe 2 Pipette 3 5 μL DI H2O onto the lower pedestal 3 Lower the arm and wait 2 3 minutes 4 Lift the arm and clean both pedestals with a new wipe Tip When thorough cleaning is required for example to remove dried sample left on the pedestals substitute 0 5M HCl for the D...

Page 285: ...venting proper formation of the liquid column when the arm is lowered The resulting spectrum may look rough or jagged If samples flatten out on the pedestal rather than beading up or forming a rounded droplet or the liquid column breaks during a measurement recondition the pedestals Supplies needed lint free laboratory wipes PR 1 pedestal reconditioning kit available from us or a local distributor...

Page 286: ... 1 compound to dry 3 Fold a clean laboratory wipe into quarters and use it to vigorously buff the surface of each pedestal Notice Support the instrument arm with one hand while you buff the upper pedestal to avoid damaging the arm Tip Black residue on the wipe is normal 4 Repeat step 3 with a new folded wipe until all residue is removed and the pedestals buff clean 5 Use canned air to remove any p...

Page 287: ...kit follow these steps 1 Lift the instrument arm and pipette 3 μL 0 5M HCl onto the lower pedestal 2 Lower the arm and wait 2 3 minutes 3 Lift the arm and clean both pedestals with a new laboratory wipe 4 Pipette 3 μL DI H2O onto the lower pedestal 5 Lower the arm and wait 2 3 minutes 6 Lift the arm and clean both pedestals with a new wipe NOTICE Support the instrument arm with one hand while you ...

Page 288: ...ntly damage the quartz fiber optic cables To prevent damage from spills keep containers of liquids away from the instrument Do not use a squirt or spray bottle on or near the instrument as liquids will flow into the instrument and may cause permanent damage Do not attempt to remove the diaphragm around the lower pedestal as it is permanently affixed to the instrument Do not allow HCl alcohol bleac...

Page 289: ... 2 3 μL diluted bleach solution see Supplies needed onto the lower pedestal 3 Lower the arm and wait 2 3 minutes 4 Lift the arm and clean both pedestals with a new wipe 5 Pipette 3 5 μL DI H2O onto the lower pedestal 6 Lower the arm and wait 2 3 minutes 7 Lift the arm and clean both pedestals with a new wipe 1 Dampen a clean soft cloth or laboratory wipe with the diluted bleach solution see Suppli...

Page 290: ...Note Clean and dry cuvettes after each measurement Use cuvettes that are free of scratches and avoid fingerprints which may affect results NOTICE Do not use a squirt or spray bottle on or near the instrument as liquids will flow into the instrument and may cause permanent damage To maintain the cuvette sampling system Keep the instrument arm closed when the instrument is not in use Use canned air ...

Page 291: ...nt to verify that throughput wavelength accuracy and bias are within specifications If the instrument has a cuvette holder NanoDrop OneC model only the test is automatically repeated using the cuvette optical path Supplies needed lint free laboratory wipes To run intensity check 1 Lift the instrument arm and clean the upper and lower pedestal with a new laboratory wipe 2 For the NanoDrop OneC mode...

Page 292: ...s to complete Here is an example of a typical intensity check result screen 6 To rerun the intensity check tap Measure 7 When finished tap End Experiment After the test is completed the results are available from the Data Viewer see example below See Manage identifiers on the instrument for details Swipe screen left to view detailed results ...

Page 293: ...d Topics Performance Verification Pedestal Image Check Performance Verification Run Performance Verification every 6 months to confirm pathlength accuracy is within specifications Supplies needed lint free laboratory wipes deionized water DI H2O calibrated precision pipettor 0 2 μL PV 1 performance verification solution liquid photometric standard available only from us or a local distributor labo...

Page 294: ...with a new laboratory wipe then pipette 1 μL DI H2O onto the lower pedestal The droplet should bead up as shown below If it does not recondition both pedestals To run performance verification 1 From the instrument home screen tap Diagnostics and then tap Performance Verification A message asks for target absorbance values Droplet beads up on properly conditioned pedestal Tap an entry box to displa...

Page 295: ... onto the lower pedestal and start the sample measurement If Auto Measure is On lower arm If Auto Measure is off lower arm and tap Measure After the measurement the software displays the results Here is an example of the performance verification result screen 7 Repeat step 6 to measure the PV 1 solution nine more times using a new 1 μL aliquot for each measurement and cleaning both pedestals after...

Page 296: ...ser Guide Thermo Scientific After each measurement a new sample result is added to the display Swipe the screen left to see a summary of the 10 sample results Swipe left again to see additional measurement details along with the overall test result Performance test result ...

Page 297: ...the PV 1 solution 9 When finished tap End Experiment and clean the pedestals with 3 5 μL DI H2O After the test is completed the results are available from the Data Viewer see example below See Manage identifiers on the instrument for details To interpret performance verification results If your instrument failed performance verification and you repeated ten measurements using 2 uL aliquots contact...

Page 298: ...n the upper and lower pedestal with a new laboratory wipe 2 Lower the arm 3 From the instrument home screen tap Diagnostics and then tap Pedestal Image Check 4 Tap Measure The instrument runs a series of tests to check pedestal position and image quality After the measurements are completed the results are displayed A green check mark indicates the instrument passed the Pedestal Image Check 5 When...

Page 299: ...hermo Scientific NanoDrop One User Guide 301 5 Safety and Operating Precautions Safety Information Operating Precautions NOTICE Be sure that all persons operating this system read the safety manual first ...

Page 300: ...The plate below the arm assembly is made of heat tempered glass The LCD display uses heat treated chemical tempered glass Both are rugged and difficult to break However should either the plate or display become cracked or broken contact us for replacement Use solvents that are compatible with the instrument see Hazardous Materials Do not use hydrofluoric acid HF on the pedestals Fluoride ions will...

Page 301: ...ymbol indicates that there is additional safety information in the documentation and failure to heed the safety precautions could result in injury WARNING Indicates a hazardous situation which if not avoided could result in death or serious injury CAUTION Indicates a hazardous situation which if not avoided could result in minor or moderate injury NOTICE Follow instructions with this label to avoi...

Page 302: ...he vicinity Only qualified persons shall perform the related procedures Avoid fire hazard Do not test flammable or explosive samples Read and follow the associated instructions carefully Avoid eye injury If you see these symbols there is a risk of exposure to ultraviolet light which can harm your eyes if safety glasses are not worn Avoid Biohazard This icon informs of a biological hazard in the ar...

Page 303: ...he protection provided by the equipment may be impaired CAUTION Avoid personal injury Perform only those procedures described in the documentation If there are other problems contact us Any other service must be performed by trained personnel CAUTION Avoid shock hazard Do not remove the cover of the instrument All service to the instrument must be performed by trained personnel NOTICE Inside the s...

Page 304: ...ock hazard Only a qualified person using the appropriate measuring device shall check the line voltage current and frequency Only our trained and certified service representatives shall attempt to service a component that carries this symbol If a protective cover on a system component appears damaged turn off the system and secure it against any unintended operation Always examine the protective c...

Page 305: ...Contact technical support for information about power conditioners and UPS Electrical Service Specifications The following table lists the specifications for electrical service Contact our service representative in your area if you have questions about the requirements Power Consumption Generally 50 more power should be available than the entire system including accessories typically uses Maximum ...

Page 306: ... housing to prevent user exposure to ultraviolet light Hazardous Materials Many standard spectroscopy methods are based on the use of solvents Others involve corrosive samples or pressurized samples in a gaseous state Volatile Solvents and Flammable Samples Compatible Solvents NOTICE Do not position the instrument so that it is difficult to operate the power switch or access the power supply and p...

Page 307: ...t to remove the diaphragm or break the seal Avoid prolonged exposure of the diaphragm to HCl alcohol bleach acetone or other solvents as the adhesive securing the seal may be affected If the seal comes loose please contact us Biohazard or Radioactive Materials and Infectious Agents Biological samples such as tissues body fluids infectious agents and blood of humans and other animals have the poten...

Page 308: ...h biohazard or radioactive materials infectious agents or any other materials and or conditions that could constitute a health or injury hazard to employees Contact us if you have questions about decontamination requirements WARNING Reduce the risk associated with potentially infectious samples Do not spill samples on any of the instrument components If spill occurs disinfect the external surfaces...

Page 309: ...countries Bluetooth is either a trademark or a registered trademark of Bluetooth Special Interest Group Windows is either a trademark or a registered trademark of Microsoft Corporation in the United States and or other countries All other trademarks are the property of Thermo Fisher Scientific inc and its subsidiaries CAUTION Indicates a hazardous situation which if not avoided could result in min...

Page 310: ...6 About this Help System 312 NanoDrop One User Guide Thermo Scientific 2015 2016 Thermo Fisher Scientific Inc All rights reserved ...

Page 311: ...55 E mail nanodrop thermofisher com Website www thermoscientific com nanodrop For International Support please contact Contact your local distributor For contact information go to http www nanodrop com Order aspx If you are experiencing an issue with your system refer to the troubleshooting information If the issue persists contact us If you are outside the U S A and Canada please contact your loc...

Page 312: ...7 Contact Technical Support 314 NanoDrop One User Guide Thermo Scientific ...

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