How to Use the NanoDrop Spectrophotometer
to Determine DNA Concentration
Wong Lab
On the Windows desktop, click the ND-1000 icon to launch the NanoDrop software. Choose
"nucleic acid measurement" from the initial program window.
1) Clean & calibrate the instrument
• Spot 2 µL of 0.5 N HCl on the sample pedestal. Lower sampling arm, and press down.
• Wipe off both pedestals with a soft Kim-Wipe (wiping multiple times is preferable).
• Spot 2 µL MilliQ-grade water onto pedestals, and clean as before.
• Load 2μl of MilliQ-grade water on the sample pedestal, and lower the upper arm.
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How to Use the NanoDrop Spectrophotometer
to Determine DNA Concentration
Wong Lab
• Type “260” in the window.
• Click "OK" in on the screen window asking you to calibrate the instrument.
• After calibration is complete, wipe both pedestals well.
2) Blank the instrument
• Spot 2μl of buffer (originally used to resuspend the DNA) onto sample pedestal.
• Lower the arm.
• Click "Blank" button on screen.
• Lift the arm and wipe both pedestals as before.
3) Measurements
• Type sample name in the “Sample ID” window on screen.
• Load 1 to 2μl of your sample on the pedestal.
• Lower the arm.
• Click "measure”. Record the concentration shown in “ng/µL” window.
• Lift the arm, and wipe both pedestals as before.
• Repeat procedure for additional samples.
4) Final results and cleaning
• When finished, click "show report", and from the new window choose "print".
• Click "exit" several times to close all windows.
• Clean the pedestals with HCl and water as in Step 1.
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.
260/230: ratio of sample absorbance at 260 and 230 nm. This is a secondary measure of nucleic acid purity.
The 260/230 values for “pure” nucleic acid are often higher than the respective 260/280 values. They are
commonly in the range of 1.8-2.2. If the ratio is
appreciably lower, this may indicate the presence of co-purified contaminants.
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