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How to measure pH of autoclaved hot agar - (Dec/11/2008 )

hello, everyone
i've been reading the methods for general and molecular microbiology from ASM press.
And in the page 310, the author mentioned that the pH of media should be measured after sterilization.

But i have a question regrading to how.

how to sterilize the pH electrode?

can i use ethanol to sterilize it?
(i'm worried about that ethanol may damage the electrode)

even if i can, i don't know how long should it be.


does anyone know

thanks alot

-Davince-

You could filter sterilize it after pH measurement

-UGA80-

I've never heard of anyone actually doing this unless they were having a problem with their cells not growing. But if I were to check the pH, I would remove a small aliquot (using sterile pipette tip) and test it using pH strips.

-smu2-

QUOTE (smu2 @ Dec 11 2008, 02:04 PM)
I've never heard of anyone actually doing this unless they were having a problem with their cells not growing. But if I were to check the pH, I would remove a small aliquot (using sterile pipette tip) and test it using pH strips.



Agree with Smu2.
Either pH strips or pH meter would do . Just need to aseptically transfer that small aliquot. Enough volume for the measurement.

-Hanming86-

QUOTE (Davince @ Dec 11 2008, 09:32 PM)
hello, everyone
i've been reading the methods for general and molecular microbiology from ASM press.
And in the page 310, the author mentioned that the pH of media should be measured after sterilization.

But i have a question regrading to how.

how to sterilize the pH electrode?

can i use ethanol to sterilize it?
(i'm worried about that ethanol may damage the electrode)

even if i can, i don't know how long should it be.


does anyone know

thanks alot


I had to do this many times during my research.

and I did it like smu2 : I took a little bit and checked the pH.
Make sure to shake the bottles before taking some out of it and let them cool down a bit too.
Do it in a sterile environment.

PS. check the pH also before autoclaving....
The pH changes during autoclaving a bit (it drops a bit).
Why? ===> If you need a certain pH after autoclaving you need to work a bit on it (adding some NaOH before autoclaving etc... after a while you know how to make the agar so that you do not always need to check the pH afterwars and need to adjust it, the more you need to do with sterile products , the more changes on infecting them.)


Do you add chloramphenicol?

-pito-

Not to put too fine a point on it, but pH is temperature sensitive. It seems to me that if the pH of the media is critical, it should be checked at the temperature at which it will be used, e.g. at 37 C.

-HomeBrew-


Thank you guys , very much.

I think i will use the universal pH indicating strip to test for it.

it was mentioned in the book that heating the media, and as carbon dioxide escapes, the pH increases
so i'm thinking about testing at 60 degrees celcius(i keep the medium in waterbath at this temp prior to pouring out)

and test the final pH as the agar solidified.

thanks, everyone:)

-Davince-

QUOTE (Davince @ Dec 13 2008, 10:11 PM)
Thank you guys , very much.

I think i will use the universal pH indicating strip to test for it.

it was mentioned in the book that heating the media, and as carbon dioxide escapes, the pH increases
so i'm thinking about testing at 60 degrees celcius(i keep the medium in waterbath at this temp prior to pouring out)

and test the final pH as the agar solidified.

thanks, everyone:)


I would test it at 37°C like homebrew said.
The most crucial phase is the autoclaving phase , thats when the pH changes.
After autoclaving, it doenst really matter a lot. The temperature then will nog influence it a lot then.



Are you sure the pH increases when heating? the pH should drop when heating the agar.
Or am I that wrong.

-pito-

Pour a test plate and let it solidify. You should test the media at ambient temperature. There are pH test probes that are made specifically for agar gel testing. Our lab just purchased one from VWR for $160. It has a standard BNC connector, fits right to our pH meter. Check it out here

-cec28007-