Mycoplasma - Mycoplasma in mammalian cultures (Dec/04/2007 )
Hello, I know this topic is covered many times but it appears fresh and new when we personally face this contamination problem.
My question: Do the cells exhibit any characteristics when contaminated with mycoplasma?
I recently grew and banked someone else's cells. I did not have any problem either while expanding or banking. They just grow excellently. But when I grew the cells in P/S free medium and sent the supernatant for mycoplasma testing, to my great shock they turned out positive!! Now I had many other cell lines running at the same time as these, so do I need to test them all? Thanks, for your replies.
-scifi-
Hi,
It pays to do routine mycoplasma testing of all your cell lines, especially if you know you have one contaminated batch.
-bob1-
QUOTE (scifi @ Dec 4 2007, 11:04 AM)
Hello, I know this topic is covered many times but it appears fresh and new when we personally face this contamination problem.
My question: Do the cells exhibit any characteristics when contaminated with mycoplasma?
I recently grew and banked someone else's cells. I did not have any problem either while expanding or banking. They just grow excellently. But when I grew the cells in P/S free medium and sent the supernatant for mycoplasma testing, to my great shock they turned out positive!! Now I had many other cell lines running at the same time as these, so do I need to test them all? Thanks, for your replies.
My question: Do the cells exhibit any characteristics when contaminated with mycoplasma?
I recently grew and banked someone else's cells. I did not have any problem either while expanding or banking. They just grow excellently. But when I grew the cells in P/S free medium and sent the supernatant for mycoplasma testing, to my great shock they turned out positive!! Now I had many other cell lines running at the same time as these, so do I need to test them all? Thanks, for your replies.
Dear "Scifi",
Cells normally exhibit no characteristics when contaminated heavily with mycoplasma's.
Yes you need to check all your cells out if they have been grown in an area where other cells have been manipulated.
WHAT TEST TO DO ?......AND WHO TO DO IT.
We use a commercial company to test our cells...please send me a private message and I will give you their details (UK company). They are experts in all things mycoplasma.
They will do the direct culture method for mycoplasma detection. In combination with Hoescht staining, these are the only FDA approved tests.
PLEASE NOTE PCR IS NOT FDA APPROVED.........The ATCC no longer offer PCR as a method for mycoplasma detection, as it is SO UNRELIABLE. We have had many instances of cells being PCR tested as negative, only to discover later they were positive all along......... YOU CANNOT PUBLISH DATA ON CELLS THAT HAVE BEEN FOUND TO BE POSITIVE.
Mycoplasma's affect everything about a cell:
Receptor Expression
Cell Metabolism
Enzyme expression
Gene Expression
Release of biologically active substances.
Induces Apoptosis
Etc Etc Etc.....the list is endless.
The world wide rate of cells infected with mycoplasma is estimated to be around 30-40%. Since introducing regular testing in my Institute, this year we have a 0% positive....i.e. the 37 cell lines sent this year have all come back negative. When I started this 12 years ago, the infection rate was 30%.
I do hope this is useful.....my only other advice to you is to quarantine ALL CELLS from every source, other than from the ATCC or ECACC.
Kindest regards
Rhombus.
-Rhombus-
Test them all.
myco usually doesn't do anything to the cells. but sometimes, it can make them act out of the ordinary.
it's better to be safe than sorry.
V
-vetticus3-
QUOTE (Rhombus @ Dec 4 2007, 02:40 PM)
QUOTE (scifi @ Dec 4 2007, 11:04 AM)
Hello, I know this topic is covered many times but it appears fresh and new when we personally face this contamination problem.
My question: Do the cells exhibit any characteristics when contaminated with mycoplasma?
I recently grew and banked someone else's cells. I did not have any problem either while expanding or banking. They just grow excellently. But when I grew the cells in P/S free medium and sent the supernatant for mycoplasma testing, to my great shock they turned out positive!! Now I had many other cell lines running at the same time as these, so do I need to test them all? Thanks, for your replies.
My question: Do the cells exhibit any characteristics when contaminated with mycoplasma?
I recently grew and banked someone else's cells. I did not have any problem either while expanding or banking. They just grow excellently. But when I grew the cells in P/S free medium and sent the supernatant for mycoplasma testing, to my great shock they turned out positive!! Now I had many other cell lines running at the same time as these, so do I need to test them all? Thanks, for your replies.
Dear "Scifi",
Cells normally exhibit no characteristics when contaminated heavily with mycoplasma's.
Yes you need to check all your cells out if they have been grown in an area where other cells have been manipulated.
WHAT TEST TO DO ?......AND WHO TO DO IT.
We use a commercial company to test our cells...please send me a private message and I will give you their details (UK company). They are experts in all things mycoplasma.
They will do the direct culture method for mycoplasma detection. In combination with Hoescht staining, these are the only FDA approved tests.
PLEASE NOTE PCR IS NOT FDA APPROVED.........The ATCC no longer offer PCR as a method for mycoplasma detection, as it is SO UNRELIABLE. We have had many instances of cells being PCR tested as negative, only to discover later they were positive all along......... YOU CANNOT PUBLISH DATA ON CELLS THAT HAVE BEEN FOUND TO BE POSITIVE.
Mycoplasma's affect everything about a cell:
Receptor Expression
Cell Metabolism
Enzyme expression
Gene Expression
Release of biologically active substances.
Induces Apoptosis
Etc Etc Etc.....the list is endless.
The world wide rate of cells infected with mycoplasma is estimated to be around 30-40%. Since introducing regular testing in my Institute, this year we have a 0% positive....i.e. the 37 cell lines sent this year have all come back negative. When I started this 12 years ago, the infection rate was 30%.
I do hope this is useful.....my only other advice to you is to quarantine ALL CELLS from every source, other than from the ATCC or ECACC.
Kindest regards
Rhombus.
Hi Thank you for both your replies. I did not know that Mycoplasma effects the cells in so many ways.
1) we do mycoplasma and IMPACT testing for all our cells from all outside sources including ATCC.
2) Our Myco testing institute is also from outside our company but within USA.
3) Regarding my other cells grown at the same time as these ones - I might have used different media bottles, but same trypsin and maybe the same hood.
4) we were thinking of curing the cells. Is this safe?
Thank you again!!!
-scifi-
General guidelines are that if a cell culture comes up positive for Mycoplasma, you're better off just chucking the culture and thawing out an old freeze of it. IF AND ONLY IF the culture is rare/impossible to get should you think about heavy doses of antibiotics to try to cure the cells of the mycoplasma infection.
-TheSquire-
QUOTE (TheSquire @ Dec 5 2007, 10:33 AM)
General guidelines are that if a cell culture comes up positive for Mycoplasma, you're better off just chucking the culture and thawing out an old freeze of it. IF AND ONLY IF the culture is rare/impossible to get should you think about heavy doses of antibiotics to try to cure the cells of the mycoplasma infection.
Hi Here, these cells were originally banked 5 yesrs ago(I wasn't here then). We are suspecting that the original bank itself is Myco +. We are in the process of testing that too and I guess, this is a rare cell line.
-scifi-
use antibiotics then.
test all cell lines in the same incubator as well.
V
-vetticus3-