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Section 13- Cell Cultures
13. Cell Cultures
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.
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Note: The most distinct difference between the NanoDrop
ND-1000 spectrophotometer "absorbance" values for cell microbial cultures
and those observed using classical cuvette based systems will be attributable to the shorter pathlength (1 mm vs. 1 cm.) Values may
not be exactly 10 fold different as readings are dependent on both the optics of a specific spectrophotometer as well as the cell type in
suspension.
The 'Cell Cultures' module displays the sample spectrum from 250 nm to 700 nm. One cursor is fixed at the frequently used wavelength
for monitoring cell suspensions (600nm) while the second cursor can be set to the wavelength of interest.
Sample Size Requirements
Field experience has indicated that 1ul samples are sufficient to ensure accurate and reproducible results when measuring aqueous
samples. However, if you are unsure about your sample composition or your pipettor accuracy, a 2 ul sample is recommended to
ensure that the liquid sample column is formed and the light path completely covered by sample.
Cell Suspension Concentrations
Due to its shorter pathlength, the ND-1000 can measure absorbencies that are 10-fold higher than those measurable on a standard
cuvette spectrophotometer. This makes it possible to directly monitor concentrated cell suspensions. Since the entire spectrum is
displayed, diluted samples exhibiting very low 'Absorbance' at 600 nm can be monitored at lower wavelengths, for example 280 nm.
Unique Screen Features
λ
600nm Absorbance: current value of the absorbance at the
1 cursor with the Baseline absorbance subtracted. Note: the actual 1
mm absorbance is displayed.
λ and Absorb.: current value of the user-selectable wavelength cursor and corresponding absorbance. The wavelength can be set by
dragging the cursor, using the up/down arrows or typing in the desired wavelength. Note: The user-selected wavelength and
absorbance are not utilized in any calculations.
Baseline: the absorbance of the user selectable-baseline (horizontal) cursor. The user may drag this cursor to a new vertical position
to create a new baseline. The absorbance value of the baseline is subtracted from the absorbance of the spectrum.
Max Absorbance: used to rescale the upper limit of the vertical axis.
Show Report: formatted for 200 samples although the buffer size can be increased to accommodate thousands of samples.
Sample Homogeneity
The user must be sure to homogeneously suspend the cells when sampling for "absorbance" measurements and read the sample
immediately to avoid significant cell settling. Vigorous mixing may be required particularly when measuring concentrated samples
13-1
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