Question

What is the Absorbance of a cell suspension? How does absorbance at 400 nm change with the number of cells in a cell suspension?

Answer 1

For bacterial cell density, we are measuring light scattering (i.e., not Absorbance) - most bacteria are colorless; therefore, the amount of light they absorb is negligible and most of the reduction in transmitted light is due to light scattering by the cells. However, if the growth media has color then it will absorb a portion of the light and interferes with your readings.

The reason most microbiologists use 600nm (red range of the spectra) for measuring ODs of the culture is:

Bacteriological media are yellow in color and will have high absorbance at 400nm but very low Absorbance at 600nm. If we are using a minimal salt media (i.e., no peptone, yeast extract...) and the growth will be spare, we can use 400nm and that will give higher absorbance.