Mycoplasma contamination - (Jun/10/2009 )

What is the best way of testing eucaryotic cell lines for Mycoplasma contamination and what is the optimal frequency of testing?

Our lab usually takes some of the cell media (1ml), centrifuges it down > take some of the supernatant and some from the bottom of the tube > run a PCR using Mycoplasma specific primers. I would say you should ideally check the media for contamination once a month. You could check more times but if you aren't experiencing problems culturing your cell lines then it might be unnecessary.

jakatta70 on Jun 10 2009, 10:08 AM said:

No no no no no no

The ATCC no longer use PCR as a method for mycoplasma detection...it is too unreliable....and these are the largest suppliers of cell lines in the world...experts

You cannot check the "media" for mycoplasma contamination ...it is an invisible enemy.

You must regularly check cell lines...you can have heavy contamination...without affecting growth or morphology....it will however affect lots of other cellular events.


GOLD STANDARD is Agar growth test in combination with Hoescht staining

All drug solutions/vaccines etc are tested this way....NOT PCR

Kindest regards

Uncle Rhombus

I´ve found couple of kits available which detect mycoplasma in culture supernatant, however, not based on PCR reaction. What about theese?

ahh - some things never change

d

Dominic on Jun 11 2009, 12:11 PM said:

Dom YOUR BACK,

thought we had lost you. Sorry for being repetitive but the same old rubbish comes onto this forum.......it's getting boring.

My question is: "Is there no proper training going on in labs anymore" ?

Kindest regards

Unlce Rhombus

rhombus on Jun 12 2009, 08:44 AM said:

Rhombus is right....

I had mycoplasma contamination last year, ran PCR and always negative. i turned to this forum and they advice me to use kits for mycoplasma detection..... they worked fine. you can use staining for mycoplasma and try to isolate in agar growth with serum and thalium acetate (BD - Agar A7). if so, mycoplasma colonies are very nice to see......

Regards

Reis, V.P.