I didn't do that myself, but a can give you some tips:

1. You will definetely nedd to keep temperature regime. 37 - or you will get not-repeatable results.

2. Do not use bicarbonate-containing bedia. Use i.e. HEPES or MOPS buffer instead, in tightly closed flasks.

3. You can make CO2-atmosphere yourself, but it is rather laborious. You will need a big empty glass jar with a cover, some vegetable oil and lots of candles. Sterilize the jar and the cover, and candles. Place candles inside the jar, light them up, place your flasks inside the jar. Close the cover (the candles should keep burning!) and seal the cover with oil to stop air exchange. If candles did fade out, it means that atmosphere inside the jar has 4.9-5.1% CO2. When you open the jar, you shold re-light the candles and reseal the cover.This receip was widely used in Russia during and after Civil War, and was told me by oldest MSU professors ))

You can use CO2 independent media such as Leibovitz-L15 media but it also depends on cell lines you are using. Most of cell lines require specific media as using different media can change rate of proliferation and cell morphology. I will strongly recommend to use C02 incubator.

In any invitro experiment optimal conditions for cell growth are essential for reliable data.

Thanks Sardar, I'm considering buying a CO2 incubator. Only that for current, I have no choice to use the only available normal incubator. I will take in mind about the media effect.

hi

You can try some device desigen by some company(Genetix biotech).

Name of device PETAKA.In this device you can grow your cell in Room temp.And by help of this you can cultue,Harvest.Transport all type of cells.

but i think u can do it with L-15 media without co2 incubator. Actually ectodermal cells are robost in nature (fibroblastic, epithelial etc). but the thing is u hve to seed them with appropriate seeding density.

I didn't do that myself, but a can give you some tips:

1. You will definetely nedd to keep temperature regime. 37 - or you will get not-repeatable results.

2. Do not use bicarbonate-containing bedia. Use i.e. HEPES or MOPS buffer instead, in tightly closed flasks.

3. You can make CO2-atmosphere yourself, but it is rather laborious. You will need a big empty glass jar with a cover, some vegetable oil and lots of candles. Sterilize the jar and the cover, and candles. Place candles inside the jar, light them up, place your flasks inside the jar. Close the cover (the candles should keep burning!) and seal the cover with oil to stop air exchange. If candles did fade out, it means that atmosphere inside the jar has 4.9-5.1% CO2. When you open the jar, you shold re-light the candles and reseal the cover.This receip was widely used in Russia during and after Civil War, and was told me by oldest MSU professors ))

What would happen if you just want to compare treatment with control sample when there is only temperature control available, can one compare the results upon treatment, secondly is there an article that defines what happen to cells, for exapmple oxidative stress, caspase activation or else...in time dependent maner without CO2 5%, normaly there is about 0.039% carbon dioxide in air! ? in case of HEK293T cells in DMEM growth medium?

Yes, make coproculture

Hi Mohamad,

If you have no incubator, its possible to had an container of CO2, It's not expensive

you can inject some little quantity of this gas in your flask, you must insert some filter to inject a sterile gas ( you must connect with tubular ) and insert an pipet on the end of the tubular. This system had an inconvenient you can't use plaque for titration…

Good luck

Apostolova,

your answer is not serious, What do you mean with coproculture, this word is french and it shows some degree of your scientific collaboration among us.

We were maintaining several cell lines in non-vent flasks without CO2 supply, and they grew well. In regular MEM (pH 6.9-7.2), without any special supplements. Ieach specific cell line will react differently on certain CO2 concentration (and hey, do all cells grow in "natural environment" at 5% CO2?) Also, the "routine" temperature fluctuations between ~35-38C did not affect cell grow significantly. So, try several cell lines that you need, and am sure you will be successful if not with all but with some of these...

Ivan, or anyone on here, could you add some context or a protocol list in how to complete mammalia cell line growth without an incubator? I will be conducting some personal research in an at home stem cell lab and am wanting to culture some mice cell lines without the finances to fund for a co2 incubator. thanks again and I appreciate all help in advance