NanoPhotometer
®
P-Class
User Manual
Version 2.1
Page 16 / 70
4.1.3
Analysis of Oligonucleotides
The procedure is as follows:
Parameter Screen
NanoVolume Applications
Cuvette Applications
Parameter Screen
Step 1
Press 1 for
NanoVolume
OR 2 for
Cuvette
folder.
Step 2
Press 1 to select
Nucleic Acids
folder.
Step 3
Press 4 to select
Oligo
mode.
Step 4
Using the NanoVolume Applications select the
Lid Factor
as
described in the “Average Detection Range Sheet” and
under 3.2.
Step 5
Enter the
Dilution Factor
using the keypad numbers. Range
1.00 to 9,999. Use the C button to backspace and clear the
last digit entered OR press
Menu/Options
to enter the
dilution factor screen. Enter the volume of the sample
using the keypad numbers. Range 0.01 to 9,999. Enter the
volume of the diluent using the keypad numbers. Range
0.01 to 9,999. Press
OK
to calculate the dilution factor
and return to the Parameters screen OR press
Cancel
to
cancel the selections and return to the Parameters screen.
Step 6
Background
correction at 320 nm is recommended to be
switched on.
Step 7
Select the
Units
of measurement using the left and right
arrows. Options: μg/ml, ng/μl, μg/μl and pmol/μl.
Step 8
Enter the
Factor
using the keypad numbers. Default value
is 33, range is 0.01 to 9,999.
Step 9
If pmol/μl is selected there are two options to set the factor
1. A selection table denoting the ratios of the 4 bases
according to the oligo sequence. Enter the proportions of
bases present using the keypad numbers and up and down
arrows to move between boxes. Default is 10 for each,
range is 0 to 9,999.
2. Enter the known extinction factor of the oligo used:
factor range 0.01 to 9,999 for ratio = [1 / extinction
coefficient *10
-6
].
Step 10
Press
OK
to enter the Results screen OR
Cancel
to
return to the
Nucleic Acids
folder.
Results Screen
Results Screen
Step 11
Apply/insert the reference sample. Press
Blank
Key. This
will be used for all subsequent samples until changed.
Step 12
Apply/insert sample and press
Sample
. This measures
at the selected wavelengths and displays the results. The
sample concentration and the ratio of A260/A280 and
A260/A230 are calculated (corrected by the background
wavelength value if selected).
Step 13
If the absorbance value of the sample is not in the linear
range a “Warning message” will pop up and “Instruction”
will be displayed in the top left corner of the result screen.
Please refer to 3.2 Software instructions/important
information on page 11 for further information.
Step 14
Repeat for all samples.
Step 15
Press
Menu/Options
to display available Options which are
described on page 8.
Step 16
Press
Escape
and confirm with
Yes
to return to the
Nucleic Acids
folder.
To change parameters, print or save methods press the Menu/Options button. The options menu will be opened. For
further explanation please see 2.3 Keypad and display on page 6 (P 300) and 7 (P330 / P 360).