User manual

Section 3- Applications
Cell Cultures
Overview
Using an absorbance spectrophotometer to monitor light scattered by non-absorbing suspended cells is common
practice in life science laboratories. Such applications, more than any other, accentuate the differences amongst the
optical systems of the numerous spectrophotometer designs.
The most distinct difference between the NanoDrop 2000/2000c pedestal “absorbance” values for microbial cell
cultures and those made with the cuvettes will be attributable to the shorter pathlength (1 mm vs. 1 cm.)
Measurements made using the pedestal mode will be approximately 10 fold lower than those made using a 10 mm
pathlength cuvette. Values between the NanoDrop 2000c and classical cuvette based spectrophotometers may not
be exactly the same as readings are dependent on both the optics of a specific spectrophotometer as well as the
cell type in suspension
.
The Cell Cultures application displays the sample spectrum from 250 nm to 700 nm.
Sample Size Requirements
Although sample size is not critical, it is essential that a liquid column is formed when using the pedestal option so
that the gap between the upper and lower measurement pedestals is bridged with sample. For cell cultures, it is
also important to ensure that the cells have not settled and that a representative aliquot has been taken of the cell
culture. It is recommended that 2 µL volumes be used when making cell culture measurements.
When using the cuvette option, it is essential that sufficient sample volume be used to ensure that the light path is
passing through a representative portion of the sample. The optical beam (2 mm) is directed 8.5 mm above the
bottom of the cuvette. Refer to the cuvette manufacturer for recommended volumes.
Sample Homogeneity
Ensure that the cell suspension is homogeneous before removing an aliquot for its “absorbance” measurement.
Vigorous mixing may be required before measuring aliquots of concentrated samples on the measurement
pedestals. Use the stirring capability when making microbial cell culture cuvette measurements.
Measurement Range
Due to its shorter pathlengths, the NanoDrop 2000/2000c can directly monitor concentrated cell suspensions. Since
the entire spectrum is displayed, dilute samples exhibiting low absorbance at 600 nm may be monitored at lower
wavelengths, for example 400 nm. Alternatively, use the cuvette mode available in the 2000c model to monitor
dilute microbial cell cultures.
Decontamination of Measurement Pedestals
If decontamination is necessary, a sanitizing solution, such as a 0.5% solution of sodium hypochlorite (1:10 dilution
of common commercial bleach solutions – freshly prepared), can be used to ensure that no biologically active
material is present on the measurement pedestals.
Note: Do not use a squirt bottle to apply the diluted bleach. Always dampen a laboratory wipe with the bleach
then clean the top and bottom pedestal surfaces as well as the outside of the instrument. Remove the
bleach solution with a lab wipe dampened with water. Do not squirt any cleaning solution into the cuvette
holder assembly as the liquid may drip into the instrument.
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