User manual

Section 3- Applications
The spectral display shows data for the current sample normalized to a 1 mm path if measured using the pedestal
mode or the pathlength selected if using the cuvette mode (model 2000c only). The pathlength information is
indicated on the Y axis.
The following features are to the right of the spectral display:
• Sample ID- field into which a sample ID is entered. The appropriate sample ID should be entered prior to
each measurement.
• Type- a drop down list from which the user may select the (color-keyed) type of nucleic acid being
measured. Options include DNA-50 for dsDNA, RNA-40 for RNA, and ssDNA-33 for single-stranded DNA.
Additional options include Oligo DNA and Oligo RNA which utilize the appropriate extinction coefficient
based upon user defined base sequences. The Custom option allows the user to enter an extinction
coefficient between 15 and 150.
• Conc- concentration based on absorbance at 260 nm and the default or user defined extinction coefficient.
Concentration units may be selected from the adjacent drop-down box. Beer’s law is used to calculate the
nucleic acid concentration as described in ‘
Nucleic Acid Calculations’.
• A260- displays absorbance at 260 nm normalized to a 10 mm path.
Note: The displayed A260 value is the difference in absorbance of the sample at 260 nm compared to the
absorbance of the sample at 750 nm. The A260 value used to calculate the nucleic acid
concentration takes into account any appropriate dye correction factors, the absorbance correction
due to the selected analysis correction nm as well as the 750 nm baseline. Therefore, the displayed
A260 value may not be the value used to calculate the nucleic acid concentration.
• 260/280- ratio of sample absorbance at 260 and 280 nm. The ratio of absorbance at 260 and 280 nm is
used to assess the purity of DNA and RNA. A ratio of ~1.8 is generally accepted as “pure” for DNA; a ratio
of ~2.0 is generally accepted as “pure” for RNA. If the ratio is appreciably lower in either case, it may
indicate the presence of protein, phenol or other contaminants that absorb strongly at or near 280 nm. See
“260/280 Ratio” in
“Diagnostic and Troubleshooting” for more details on factors that can affect this ratio.
• Dye 1 (or 2): user selected dye(s). The default selections are Cy3 for Dye 1 and Cy5 for Dye 2.
- Abs - the normalized absorbance of the respective dye at the 1 mm pathlength.
- pmol/ µL - is the concentration based upon the respective dye’s extinction coefficient. Concentration
units may be selected from the adjacent drop-down list box.
• Analysis correction - the absorbance at the specified correction wavelength is subtracted from the
absorbance at the analysis wavelength before calculating the concentration. This only affects the reported
nucleic acid concentration. The default analysis wavelength is 340 nm.
Note: The software normalizes the visual spectrum display for all readings at 750 nm and will
automatically calculate a baseline between 400 nm and 750 nm for dye concentration
calculations.
Making Micro Array Measurements
1. Select the Micro Array application from the main menu. If the wavelength verification window appears,
ensure the arm is down and click OK.
2. Select the type of sample to be measured from the Type drop-down list. The default setting is ssDNA-33.
3. Choose the concentration units from the drop-down list adjacent to the color coded concentration box. The
default units are ng/µL.
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