14 replies to this topic

[Fiaq Khan]

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

[pito]

Fiaq Khan, on 25 April 2013 - 04:32 AM, said:

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

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...

[Paulgs3]

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.

[pito]

Paulgs3, on 26 April 2013 - 04:26 AM, said:

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.

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.

[Fiaq Khan]

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?

Edited by Fiaq Khan, 27 April 2013 - 03:09 AM.

[Paulgs3]

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

[pito]

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

[Phil Geis]

USP (US Pharmacopeia) recognizes standard deviation of plate counts to be 0.5 log.  How many plates were inoculated?
http://www.microbiol..._.2011.17_3.pdf

[Phil Geis]

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.

[lyok]

Phil Geis, on 01 May 2013 - 04:34 AM, said:

USP (US Pharmacopeia) recognizes standard deviation of plate counts to be 0.5 log.  How many plates were inoculated?
http://www.microbiol..._.2011.17_3.pdf

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.

[Phil Geis]

It addresses only the method's capability.

[lyok]

Phil Geis, on 04 May 2013 - 08:33 AM, said:

It addresses only the method's capability.

Can you give a bit more explenation?
I am not sure if I understand you.

I also dont understand how you get that "standard deviation plate counts to be 0.5 log" from the paper you attached.

If I understand you correct you mean that the 0,5 log deviation is adressed to the method, so you mean that plating bacteria always has a 0,5 log deviation because of the plating?
And you do mean 10^-0,5 X + 0,5 , right? A half log difference?

[Phil Geis]

Right.

[lyok]

Phil Geis, on 08 May 2013 - 06:39 AM, said:

Right.

Ok I see what you mean!
Thanks

[memari]

A colony origins from one  to more than 100 bacteria.
Use a inoculating turntable, and an Ultrasonic bath(Jewelry Cleaner) to disociate bateria from each other.