CFU measurments - (Apr/25/2013 )

I have done CFU measurement of E.coli at OD~0.3. I am getting different results as describe below:

First time I did the experiments the CFU results showed that 0.3 OD = 2.07 x 10⁸ cells/ml.

After some weeks I did the same experiment and the CFU results are showed to me that 0.3 OD = 1.41 x 10⁹ cells/ml.


I am not sure why I am getting that different values of CFU for same OD?

Any help will be highly appreciated. Thanks

Fiaq Khan on Thu Apr 25 12:32:23 2013 said:

Did you do everything the same way? Same protocol?
It can be because of so many things... Not the same amount of cells at the start... not the same time of incubation? Less or more aeration....
Also: in the end the difference is not that big...

I agree there could be a ton of reasons why: viability caused by incubation time, media formulation, incubation temp, shaker speed....

A log difference in cfu is rather significant to me.

Paulgs3 on Fri Apr 26 12:26:58 2013 said:

if the samples where the same yes.
But here: since we dont know what might have gone "wrong" or different it means pretty much nothing.
Maybe I was not clear enough about that.

I have done everything same. Same bacteria, same media and same OD which is 0.3. Is that difference in CFU is acceptable? or how much variation is possible?

If it was me, I'd gather more than two data points.

It also depends on why you need this data...

USP (US Pharmacopeia) recognizes standard deviation of plate counts to be 0.5 log. How many plates were inoculated?
http://www.microbiol.org/wp-content/uploads/2010/07/Sutton.jvt_.2011.17_3.pdf

btw - did you generate a standard curve that includes these bacterial titers - or were these just 2 isolated observations? You should ensure that you can (by standard curve) differentiate between such high concentrations of cells.

Phil Geis on Wed May 1 12:34:52 2013 said:

How do you get that 0,5 log?
I see a standard deviation of 10-15%.

you do mean half a log difference?
Meaning 2.10^4 bacteria would still be the same as 2.10^3,5 bacteria? It seems a bit weird, because its 20000 bacteria compared with 6324.