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CO2 incubator - transfected clones - will they survive? (Nov/15/2006 )

hai all
am really very unlucky. i just have my transfected clones growing up and now, my CO2 incubator is not working. it is not taking in CO2. what will happen to my clones now?
i think it will take a minimum of two days for ir to be serviced. i dont have any spare incubaotrs available? will these cells survive?


I can't answer your question but I can give you advice I got in same situation - you can produce CO2 in desired amount to get right concentration in an incubator.
eg. sodium carbonate + citric acid -> sodium citrate + CO2 + H2O
Just fix all stechiometry and calculate right amount of reagents to get as much of CO2 as you need (1 mol of gas is 22,4dm3 at standard conditions).


Here are a few ideas for you:
1. Purge the air in the culture flask with sterile 95%Air/5%CO2. You can use a sterile 0.2 um Acrodisc filter then cap the flask tight. People frequently do this with roller bottle cultures. You'll be better off if your cell density is lower. You can generate the 95% Air/5%CO2 using rotometers off a bioreactor.
2. Another idea is to use HEPES in the medium, say about 24 mM. You'll need to prepare bicarbonate free medium then you can put the flask in a non-CO2 incubator. Leibovitz L-15 is an example of a bicarbonate/non-CO2 requiring media. A PubMed search for this media will give you some more background.


hai can u please tell me how to adjust the reaction to generate 5% CO2.


you may adjust pH with Hepes too.


QUOTE (bioenthu @ Nov 16 2006, 01:39 AM)
hai can u please tell me how to adjust the reaction to generate 5% CO2.

If you have two rotometers you can regulate the flow rate to 95% Air and 5% CO2. An easy options is to use the controls on a bioreactor and then sterile filter the mixed 95%Air/5%CO2


Probably this reply is too late. Don't worry if it is a question of just two days. Change cell culture medium every day untill you have your incubator repaired or just use Leibovitz L-15 medium.