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Measuring OD

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Measuring OD

Postby pab94055 » Fri Dec 10, 2010 1:36 am

Hello,

I want to measure the OD of various bacteria. the plate reader I have can only read at OD595. Will that be ok if its not exactly OD600 as the literature suggest?

Thanks
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Postby JackBean » Fri Dec 10, 2010 6:40 am

I would try to measure full spectrum first, but I think, it should be OK (although you will have smaller optical density!).
http://www.biolib.cz/en/main/

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Postby canalon » Fri Dec 10, 2010 4:11 pm

Actually the wave length does not matter at all.
When you are measuring cell density with a spectrophotometer, you are not really measuring the absorption of light, but its scattering by the particles (cells). And it is the same at any wavelength. The only thing that you might want to recheck is the equivalence OD=number of cells, as the values ate different wavelength will vary with many parameter (but that should be true even at 600nm, because the geometry of the different species of bacteria will influence the equivalence between OD and cell numbers). Yet honestly I am ready to bet that there will be no difference between the 2 measurements.
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Postby jwalin » Sat Dec 11, 2010 4:27 pm

hmm that is an interesting point...
hmm wait but isnt OD equivalent to absorbance?
it isn't what you do that matters but it is how you do it
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Re:

Postby canalon » Mon Dec 13, 2010 4:44 pm

jwalin wrote:hmm that is an interesting point...
hmm wait but isnt OD equivalent to absorbance?


No they are not. The bacteria are not absorbing the light. You are truly measuring absorbance when you measure concentration of molecules in solutions, like DNA (260nm) or proteins or pigment. But for particles like bacteria, it is just the scattering of the light (% transmission) that is measured, so the wave length is not really important as long as the blank is correct.
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Postby JackBean » Tue Jan 04, 2011 4:13 pm

canalon: it's not exactly the same, I think up to ~300 (maybe 400), there is no OD, but then it grows up.
http://www.biolib.cz/en/main/

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