One review ( Article Lactobacilli and pediococci as versatile cell factories – Ev...
) mentioned that "Many LAB are known to produce exopolysaccharides (EPS), and care should be taken with interpreting OD-values as these will be influenced by the produced EPS" and I noticed some papers showed they measure the OD at 595 nm instead of 600 nm. Could someone explain thoroughly why people choose 595 nm but not other wavelength?
Hi Xiangang
It does not seem to be logically necessary to choose exactly 595nm. For optical densities you are measuring a transmission in a turbid solution not an extinction.
You avoid wavelenghts close to the UV spectra to filter out results from dissolved molecules and blank against your medium to eliminate VIS-Colours in the samples
If there is a possible maximum extinction of undissolved by-molecules that have maximum of exactly 595nm, they will be already visible at 590nm or 600nm.
You have to take in mind too, that a linear progression of the results for turbid solutions is only given up to arround ext. 0,7, above this value it is usefull to dilute.
Possible reasons might also be: Perhaps researchers had good experiences with this wavelenght.. or used instruments with exactly this (fixed) filter.
Anyway: the value for ODs can differ between single photometers it is not 100% consistent, if you want to exactly estimate the rate of living cells, you should perform a living cell count, f.e. via plating.
For bacterial cell density, we meausre light scattering and not Absorbance. Since most of the bacteria are colorless, hence, the amount of light absrbed by them is negligible and most of reduction in transmitted light takes place due to scattering of lights because of cells.
A wave length anywhere between 400nm and 700nm fir measurement of optical density can be used. But, the amount of light scattered is proportional to the ratio of the cell (particle) size to the wavelength of the incident light. Or we can say that the light scattering exhibited by same number of cells per mL will be higher at 400nm as compared to 600.
The main reason for usage of red range of spectra for measurement of OD is that color of the bacterial growth media is yellowish and has a higher absorbance at 400nm while at 600nm, the absorbance is very low.hbb
why would ou need to use UV? why not CFU count? more reliable. UV may detect cell debres and dead cells etc and may effect your observation of live bacterial mass. i prefer CFU instead of UV for microbes related wokr not only LAB.
Hi Xiangang,
I found the response by Frank Hoger quite complete.
I would just like to reinforce the part that says "Perhaps researchers had good experiences with this wavelenght.. or used instruments with exactly this (fixed) filter" as the probable cause for researchers to adopt 595 nm instead of 600 nm.
BW,
Juliano
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