detection of mycoplasma with hoechst staining - (Oct/20/2010 )

Hello
I would like to stain some cancer cell line with hoechst to detect an eventual mycoplasma contamination.
Any protocol I find suggests to use Hoechst 33258, in our lab we only got Hoechst 33342: would it be good anyway?
what is the difference between the two of them? do they need just different filters to be observed with the fluorescent microscope or do they bind differently to the Dna?
thank you for your advice!

Hi,

The key difference between them is that the additional ethyl group of Hoechst 33342 renders it more lipophilic, and thus more able to cross intact cell membranes. In some applications, Hoechst 33258 is significantly less permeant.
With reagrd to the mycoplasma infection: I would recommend a PCR based mycoplasma detection because it is much more sensitive. With Hoechst you will only see if your cells are really severly infected.

regards