How much serum should I put in the tissue culture - (Oct/11/2007 )
Hello,
Recently I have established a immortalized cell line from mouse embryonic fibroblast. However, at certain stage, after selection of GFP expressing cells, I lost a lot of cell and they grow unbelievably slow. Therefore, I switched to 20% serum medium to rescue the cells and expand the cell line. Now they grow perfectly fine. My question is, should I continue to use 20% serum or I should go back to lower concentration?
Will it harm the cells if I continue to use 20% serum?
Thanks
I don't think the high serum levels will hurt your cells but it will work out to be pretty expensive long-term to keep using that much serum. Have a look at this website, it might help you to cut back on your serum levels.
Good luck with it!
http://www.corning.com/Lifesciences/techni..._cls_an_074.pdf
Good luck with it!
http://www.corning.com/Lifesciences/techni..._cls_an_074.pdf
Hey, Macgirl, thank you for your information.
you could always just have slower growing cells - i'm working on rho zero's and i havn't had to split them all week - makes life much easier - are there any suppliments you could add rather than the whole sera?
dom
5~20% is good. but if 20% works for your cells, why not? (except that it is gonna be expensive). maybe you can split your cells to 1:4 in 96 wells and do a duplicate/triplicate testing on 2,5,10,15,20,25% serum and see what works best within a week.
Recently I have established a immortalized cell line from mouse embryonic fibroblast. However, at certain stage, after selection of GFP expressing cells, I lost a lot of cell and they grow unbelievably slow. Therefore, I switched to 20% serum medium to rescue the cells and expand the cell line. Now they grow perfectly fine. My question is, should I continue to use 20% serum or I should go back to lower concentration?
Will it harm the cells if I continue to use 20% serum?
Thanks
Dear Ching,
In all your replies so far no one has mentioned the QUALITY of the serum ?
With some serum you may have to use 20%. However if you use quality serum you may still get away with 7.5%-10%.
For the users of serum, this is the rank order for serum quality.
New Zealand>Australian>South American>North American>European>British.
In our lab we have proven this rank order over the past 20 years. We batch test all our serum in order to get Accurate and Precise experimental data. The following markers are tested:
Cell Growth
Viabilty
Induction of Enzymes
Concentration of cGMP in smooth muscle cells
Expression of sodium channels in transfected HEK cells
Concentration of Cytochrome C in Jurkat cells
Oxygen consumption in J774 and Jurkat cells
Oxidation/Reduction states of mitochondrial enzyme complexes (I-IV).
I would recommend to all experimenters to BATCH TEST their serum and always if possible go for NEW ZEALAND SERUM.
Kindest regards
Rhombus