TITLE: Nanodrop
AUTHORS: Allyson Hirsch, George  Testo

[DESCRIPTION]
The Thermo Scientific NanoDrop 8000 Spectrophotometer is a full-spectrum (220-750nm) instrument that accurately measures up to 8 individual 1 ul samples in one measurement cycle. The software allows the user to measure samples using either the full 8-position mode or a convenient single sample mode. The NanoDrop 8000 utilizes the same patented sample retention technology employed on all NanoDrop™ instruments. The surface retention system holds the sample in place eliminating the need for cumbersome cuvettes and other sample containment devices. Clean-up is accomplished in seconds. In addition, the NanoDrop 8000 has the capability to measure highly concentrated samples without dilution (50X higher concentration than the samples measured by a standard cuvette spectrophotometer).

[GUIDELINES]
Up to eight 1 ul samples are pipetted onto the sample pedestal using a low volume multi-channel pipettor.  Each position is actually the end of a fiber optic cable (the receiving fibers). A second set of fiber optic cables (the source fibers) are brought into contact with the liquid samples causing the liquid to bridge the gaps between the fiber optic ends. The pathlengths are automatically controlled to 1mm and 0.2 mm paths. Readings are acquired through sequential measurement across the 8 positions. A pulsed xenon flash lamp provides the light source and a spectrometer utilizing a linear CCD array is used to analyze the light that passes through the samples. The instrument is controlled by PC based software, and the data is logged in an archive file on the PC.

[STEPS]
SECTION: Preparing &amp; Blanking Nanodrop
1. Allow samples to efficiently thaw before starting (10 min)

SECTION: Preparing &amp; Blanking Nanodrop
2. Log into computer and open the Nanodrop program and select “Nucleic Acid."
SECTION: Preparing &amp; Blanking Nanodrop
3. Clean Nanodrop pedestals using molecular grade H2O.
 

SECTION: Preparing &amp; Blanking Nanodrop
4. Follow program instructions & load 2 µL of molecular grade H2O onto each sample pedestal.

SECTION: Preparing &amp; Blanking Nanodrop
5. Blank, then immediately clean sample reader pedestals with molecular grade H2O and a new Kimwipe.
SECTION: Preparing &amp; Blanking Nanodrop
6. Add 2 µL of blanking solution (solvent used to reconstitute nucleic acids).

 

SECTION: Preparing &amp; Blanking Nanodrop
7. Blank the machine and clean sample reader pedestals immediately with molecular grade water and a new Kimwipe.
SECTION: Preparing &amp; Blanking Nanodrop
8. Label your samples in the program: can either be pre-programmed or manually entered...

SECTION: Preparing &amp; Blanking Nanodrop
9. Make sure the correct sample desiring to be read is selected: DNA or RNA.
SECTION: Loading Samples onto Nanodrop
10. Load 2 µL of sample into the correct sample reader pedestals and click “Measure.”

SECTION: Loading Samples onto Nanodrop
11. Clean with molecular grade H2O and a new Kimwipe after each run.
SECTION: Loading Samples onto Nanodrop
12. Repeat steps 10-11 until all samples have been quantified using Nanodrop.
SECTION: Troubleshooting DNA/RNA Concentrations
12.1. To determine if samples need a re-run:
DNA/RNA concentrations less than 10ng/uL
A 260/280 ratio outside of 1.8-2.0 for DNA:
A 260/280 ratio above 2.0 for RNA
 

SECTION: Cleaning up Sample Pedestals
13. Clean sample reader pedestals after last use with molecular grade H2O and a new Kimwipe; make sure to clean and lower the sample pedestal arm after use.
SECTION: Cleaning up Sample Pedestals
14. Save the Nanodrop table information to its correct location.