background image

UltiMate 3000 Nano and Cap - System Installation and Application 

 

39

3.6 Parallel LC 

This application allows you to increase the throughput for peptide separations. While the 
autosampler's injection valve is in 

Load

 position, sample is drawn by the syringe through the 

needle and placed in the sample loop. The needle pair withdraws liquid from the well or vial 
and then descends into the wash port. The injection valve is switched to the 

Inject 

position, 

allowing the loading pump to direct the sample from the sample loop onto trap column 1 (in 
the FLM-3200; 

 Fig. 21, no. 9) for sample preconcentration. (For information about how to 

install the trap columns, refer to section 3.7.) 

Valve 1 (left switching valve in the FLM-3200) is in position 

10-1

. In parallel, valve 2 (in the 

FLM-3300) is in position 

1-2 

to allow the wash step to elute remaining hydrophobic compounds 

from trap column 2 (in the FLM-3300; 

 Fig. 21, no. 11) and separation column 2 (in the 

FLM-3300; 

 Fig. 21, no. 19). The nano valve (right switching valve in the FLM-3200) is in 

position 

6-1

 to direct the eluent from the wash-step to the detector. 

When the sample is loaded on trap column 1, valve 1 is switched to position 

1-2

. The analytical 

pump 1 then directs the sample from trap column 1 to separation column 1 (in the FLM-3200; 

 Fig. 21, no. 14). At the time when the first peaks are expected, the nano valve is switched to 

position 

1-2

 to direct the eluent to the detector. In the meantime, separation column 2 is 

reconditioned. Shortly before the separation ends, valve 2 is switched to position 

10-1

. Trap 

column 2 is conditioned with the loading solvent. 

As soon as the gradient over separation column 1 is finished, a new sample is injected. Valve 1 is 
in position 

1-2. 

Valve 2 is in position 

10-1

 to load the sample on trap column 2. At the same time, 

separation column 1 is washed and its eluent is collected, with the nano valve being in position 

1-

2

.  

When the trap column 2 is loaded, valve 2 is switched to position 

1-2

. This connects the 

separation column 2 with the trap column 2 (analytical pump). At the end of the wash step for 
separation column 1, the nano valve is switched to position 

6-1

. The separation column 1 is 

reconditioned while the separation runs on separation column 2. Before the experiment ends, 
valve 1 is switched to position 

10-1

 to prepare for the next injection. 

This sequence, which can be repeated as often as needed, allows running the analysis on one 
column and simultaneously washing and equilibrating the other column. 

  

Please note:

  The application comprises two pumps (LPG-3400 and DGP-3600) and two 

flow managers (FLM-3200 and FLM-3300; 

 Fig. 20, page 40). 

 

Therefore, adapt the names under which the devices are identified in the 
installation environment and in the Chromeleon client in the Chromeleon 
Server Configuration program. Keep in mind that you may have to adapt 
the links to the associated control panels as well. 

Summary of Contents for UltiMate 3000

Page 1: ...UltiMate 3000 Nano and Cap System Installation and Application Revision 1 0 Date May 2006 2006 Dionex...

Page 2: ...UltiMate 3000 Nano and Cap System Installation and Application...

Page 3: ...guration 16 3 2 Direct Injection 20 3 3 Preconcentration 24 3 4 2D LC Salt Plug 28 3 5 Comprehensive 2D LC 33 3 6 Parallel LC 39 3 7 Installing a Trap Column 45 4 Automated Control by Chromeleon 47 4...

Page 4: ...Salt Plug Kits 65 8 4 1 Standard Devices 65 8 4 2 Biocompatible Devices 65 8 5 Comprehensive 2D LC Kits 65 8 5 1 Standard Devices 65 8 5 2 Biocompatible Devices 65 8 6 Parallel LC Kits 66 8 6 1 Stand...

Page 5: ...lt in damage to the instrument Important Indique que ne pas tenir compte de l information jointe peut endommager l instrument Warning Indicates that failure to take note of the accompanying informatio...

Page 6: ...te the special properties of the solvents such as viscosity boiling point UV absorption UV VIS detector and refractive index refractive index detector Buffer concentration Typically up to 1 mol L 1 3...

Page 7: ...ilable system modules refer to the table below If you have any questions do not hesitate to contact your Dionex sales representative or distributor Please note For more information about the configura...

Page 8: ...th two 10 port micro switching valves Standard flow manager FLM 3100 Biocompatible flow manager FLM 3100B or FLM 3200 B with 10 port micro and 6 port nano switching valves Standard flow manager FLM 32...

Page 9: ...WPS 3000 B Autosampler without temperature control Standard autosampler WPS 3000 Biocompatible autosampler WPS 3000B 5820 0010 5821 0010 5820 0020 5821 0020 1 5 Intended Use The UltiMate 3000 system i...

Page 10: ...UltiMate 3000 Nano and Cap System Installation and Application 6...

Page 11: ...l the condensation is completely gone Install the system in the laboratory on a stable surface that is free of vibration Make sure that the surface is resistant to solvents Avoid locations with extrem...

Page 12: ...nstallation and Application 8 2 2 Rear Panel Connectors Fig 2 Rear panel view of an UltiMate 3000 system Please note Fig 2 illustrates how the system modules are interconnected This applies accordingl...

Page 13: ...to the Chromeleon server PC via only one USB connection e g from the autosampler Fig 2 Please note The SRD 3600 solvent rack can be operated with Chromeleon via the pump For information about the conn...

Page 14: ...s Observe the information about the facility requirements when installing the system Connect the drain tubing from the devices accessories kits to the outlet of the modules to direct liquids that migh...

Page 15: ...e of the flow manager separately to the waste container Always connect the drain tubing from the bottom module as shown in Fig 4 c a b c Fig 4 Connected drain tubing Important The drain tube must rema...

Page 16: ...ent Connect the modules on the rear panel as described in Rear Panel Connectors page 8 For examples of the fluid connections between the system modules refer to Application Examples and Fluid Connecti...

Page 17: ...es raccords uniquement dans aux endroits o ils sont pr vus Important When you operate biocompatible system modules observe the instructions for connecting the capillaries and other components Refer to...

Page 18: ...and ferrules only for the same capillary connection This is to avoid increased dead volume Important La r utilisation des raccords et ferrules n est possible que pour la connexion capillaire d origin...

Page 19: ...cations split ratio 1 100 6721 3160A Splitter cartridge biocompatible for capillary HPLC applications split ratio 1 15 6721 3165A Splitter cartridge biocompatible for micro HPLC applications split rat...

Page 20: ...ludes the modules listed in the table below In addition the appropriate Basic Configuration Kit section 8 1 page 64 is required Part No System Module Nano HPLC Capillary HPLC Micro HPLC Monolith HPLC...

Page 21: ...nd Application 17 Fig 6 and the related table provide an overview of the fluid connections for manual injection For an example of the fluid connections for nano HPLC also refer to Fig 7 page 19 Fig 6...

Page 22: ...le 8 Capillary from switching valve to flow splitter both in flow manager Standard Biocompatible 6720 00332 6721 00332 6720 00342 6721 00342 6720 00352 6721 00352 6720 00342 6721 00342 9 Capillary fro...

Page 23: ...UltiMate 3000 Nano and Cap System Installation and Application 19 The part numbers in Fig 7 refer to nano HPLC standard devices Fig 7 Fluid connections for manual injection nano HPLC standard devices...

Page 24: ...s listed in the table below In addition the appropriate Direction Injection Kit section 8 2 page 64 is required Part No System Module Nano HPLC Capillary HPLC Micro HPLC Monolith HPLC SR 3000 5035 920...

Page 25: ...nd Application 21 Fig 9 and the related table provide an overview of the fluid connections for direct injection For an example of the fluid connections for nano HPLC also refer to Fig 10 page 23 Fig 9...

Page 26: ...r 6035 25502 standard or 6037 25502 biocompatible 6035 25532 standard or 6037 25532 biocompatible 8 Capillary from separation column to detector flow cell The flow cell is shipped with appropriate cap...

Page 27: ...ltiMate 3000 Nano and Cap System Installation and Application 23 The part numbers in Fig 10 refer to nano HPLC standard devices Fig 10 Fluid connections for direct injection nano HPLC standard devices...

Page 28: ...ou stack the system modules as shown below Fig 11 Module arrangement for preconcentration The system includes the modules listed in the table below In addition the appropriate Preconcentration Kit sec...

Page 29: ...d Application 25 Fig 12 and the related table provide an overview of the fluid connections for preconcentration For an example of the fluid connections for nano HPLC also refer to Fig 13 page 27 Fig 1...

Page 30: ...w manager switching valve Standard Biocompatible 6720 00332 6721 00332 6720 00342 6721 00342 6720 00342 6721 00342 9 Trap column holder with the appropriate capillaries and Trap column 6720 00122 1604...

Page 31: ...ltiMate 3000 Nano and Cap System Installation and Application 27 The part numbers in Fig 13 refer to nano HPLC standard devices Fig 13 Fluid connections for preconcentration nano HPLC standard devices...

Page 32: ...nished the ion exchange column is removed from the fluidic path by switching the right valve to position 1 2 This allows desalting the sample collected on the trap column with the loading pump After t...

Page 33: ...3600 5035 9230 DGP 3600 5035 0050 standard or 5037 0060 biocompatible VWD 3100 VWD 3400 5074 0005 5074 0010 Please note that flow cells must be ordered separately for order numbers see page 15 FLM 31...

Page 34: ...d Cap System Installation and Application 30 Fig 15 and the related table provide an overview of the fluid connections for 2D LC salt plug Also refer to Fig 16 page 32 Fig 15 Fluid connections for 2D...

Page 35: ...332 standard or 6021 0033 biocompatible 9 Trap column holder with appropriate capillaries and Trap column 6720 00122 1604542 10 Separation column The column is connected directly to the switching valv...

Page 36: ...m Installation and Application 32 Fig 16 too illustrates the fluid connections for 2D LC salt plug The part numbers in the picture refer to the standard devices Fig 16 Fluid connections for 2D LC salt...

Page 37: ...eluting from the ion exchange column are trapped on trap column 2 collecting a new peptides fraction At the end of the separation on the reverse phase column valve 1 is switched to position 1 2 to co...

Page 38: ...820 0010 standard or 5821 0010 biocompatible 5820 0020 standard or 5821 0020 biocompatible SRD 3600 5035 9230 DGP 3600 5035 0050 standard or 5037 0060 biocompatible VWD 3100 VWD 3400 5074 0005 5074 00...

Page 39: ...tion 35 Fig 18 and the related table provide an overview of the fluid connections for 2D LC comprehensive for capillary 1st dimension nano 2nd dimension HPLC Also refer to Fig 19 page 38 Fig 18 Fluid...

Page 40: ...flow splitter cap or micro 6720 00282 standard or 6721 0028 biocompatible 9 Capillary from autosampler to ion exchange column SCX 6720 00582 standard or 6721 00582 biocompatible 10 Ion exchange column...

Page 41: ...35 25542 standard or 6037 2554 biocompatible 20 Capillary from nano flow splitter to switching valve 6720 00332 standard or 6021 0033 biocompatible 21 Sample loop 20 l 6820 00182 standard or 6821 0018...

Page 42: ...ion 38 Fig 19 too illustrates the fluid connections for comprehensive 2D LC for capillary 1st dimension and nano 2nd dimension HLPC The part numbers in the picture refer to the standard devices Fig 19...

Page 43: ...no valve is switched to position 1 2 to direct the eluent to the detector In the meantime separation column 2 is reconditioned Shortly before the separation ends valve 2 is switched to position 10 1 T...

Page 44: ...21 0010 biocompatible 5820 0020 standard or 5821 0020 biocompatible SRD 3600 5035 9230 DGP 3600 5035 0050 standard or 5037 0060 biocompatible VWD 3100 VWD 3400 5074 0005 5074 0010 Please note that flo...

Page 45: ...on and Application 41 Fig 21 and the related table provide an overview of the fluid connections for parallel LC For an example of the fluid connections for nano HPLC also refer to Fig 22 page 44 Fig 2...

Page 46: ...opriate capillaries and Trap column 6720 00122 1604542 6720 00122 1604542 1649722 10 PEEKsil bridge from left switching valve FLM 3200 to FLM 3300 switching valve 6720 00592 standard or 6721 00592 bio...

Page 47: ...332 6721 00332 6720 00342 6721 00342 6720 00342 6721 00342 19 Separation column The column is connected directly to the switching valve 1603212 1602952 164092 20 Connection of separation column FLM 33...

Page 48: ...UltiMate 3000 Nano and Cap System Installation and Application 44 The part numbers in Fig 22 refer to nano HPLC standard devices Fig 22 Fluid connections for parallel LC nano HPLC standard devices...

Page 49: ...ne see Fig 23 The retention times with known sample shift to later elution The column pressure drop is larger than 10 bar Please note The pressure stability option of the UltiMate 3000 system Hold pre...

Page 50: ...a trap column cartridge in the holder 2 Slide the ferrule of each pre assembled connecting tubing approximately 3 mm 0 1 inch back on the tubing to extend the length of the tubing that enters the trap...

Page 51: ...the USB cables that are shipped with the modules Dionex recommends connecting all modules to one central module e g the autosampler and connecting this module to the Chromeleon Server PC Fig 2 page 8...

Page 52: ...Preconcentration or 2D LC Salt Plug Please note The Operating Instructions for the modules also provide an overview of the different commands and properties supported by Chromeleon for the respective...

Page 53: ...nly if both the pump and the flow manager are installed in Chromeleon The default entry is None To change the setting click the arrow next to the uses FLM 3x00 Flow Splitter input field and select the...

Page 54: ...the master pump i e the total flow which is the flow before the flow splitter Pressure and or MasterPressure Indicates the pressure Pumps without flow splitter assignment Pressure indicates the pump p...

Page 55: ...default name ColumnPressure under which the column pressure signal is identified in the installation environment When your system includes more than one flow manger Adapt the name for all additional f...

Page 56: ...ent signal value read only UpperLimit sets the upper signal limit and LowerLimit sets the lower signal limit AcqOn starts data acquisition AcqOff terminates data acquisition Retention reports the rete...

Page 57: ...phase of the pump cycle On the Segments Pump Link tab page of the autosampler s properties determine which synchronization is performed Please note Dionex recommends always enabling synchronization a...

Page 58: ...ese properties from the Commands dialog box does not change the standard synchronization setting on the Segments tab page For more information refer to the Operating Instructions for the autosampler a...

Page 59: ...to change certain settings and or perform certain commands A magnetic Dionex menu pen part no 6300 0100 is included in each module s accessories kit White spots on the front panel mark the positions o...

Page 60: ...UltiMate 3000 Nano and Cap System Installation and Application 56...

Page 61: ...PC Check the connection to the server PC The USB port on the computer is not ready for operation Check the USB port on the computer The system has very high backpressure One or more capillaries in th...

Page 62: ...mn The solvent is degraded Use fresh solvent The flow splitter is clogged or defective Replace the splitter cartridge Flow Manager manual If this does not solve the problem contact Dionex Service Tria...

Page 63: ...n appropriate solvent No flow The system is leaking Find and eliminate the leak The check valves are not installed properly in the pump not in direction of flow or defective Correctly install or repla...

Page 64: ...The degasser channels are contaminated Rinse the degasser channels Solvent Rack manual The solvents are degraded or dirty or their purity is insufficient Use fresh solvents Contamination occurs somewh...

Page 65: ...idual instruments provide instructions for shutting down the instrument and describe all maintenance procedures that can be carried out by the user Dionex personnel should perform any additional servi...

Page 66: ...UltiMate 3000 Nano and Cap System Installation and Application 62...

Page 67: ...patible with the parts they replace Please note The part numbers always refer to the packing unit Unless stated otherwise the packing unit is one unit Please note For more information about which acce...

Page 68: ...on Kit biocompatible for Capillary HPLC 6721 0082 Basic Configuration Kit biocompatible for Micro HPLC 6721 0083 Basic Configuration Kit biocompatible for monolithic HPLC 6721 0084 8 2 Direct Injectio...

Page 69: ...llary HPLC 6721 0048 Preconcentration Kit biocompatible for Monolithic HPLC 6721 0049 8 4 2D LC Salt Plug Kits The kits include all parts that are necessary for the configuration 8 4 1 Standard Device...

Page 70: ...0055 Parallel LC Kit biocompatible 2x Monolithic 6721 0056 8 7 Upgrade Kits 8 7 1 Standard Devices Description Part No Upgrade CAP LC Kit with flow cell for UVD 3000 6720 0094 Upgrade CAP LC Kit with...

Page 71: ...ngths User input and display Large LCDs programmable to show important system parameters Soft keys for operation during installation and maintenance System control All functions and parameters are sof...

Page 72: ...UltiMate 3000 Nano and Cap System Installation and Application 68...

Page 73: ...8 D Degasser 54 Degasser Parameters 54 DegasserVacuum 54 Direct Injection 20 64 E Error Messages Chromeleon 60 F Facility Requirements 7 Flow Splitter 49 Fluid Connections 13 Front Panel 9 I Injectio...

Reviews: