Hi all,

Is there anyone who has experience using EBV B cells PLEASE?

We initially received the cells from a collaborator and were able to grow them and prepare freezing stocks in liquid Nitrogen. But now that we take vials out of liquid Nitrogen to grow for experiments, they are okay for the 1st few hours and then they start developing these small feature/ cells.

As the cells were sourced from another research institute, I sent them out for Mycoplasma test and the results were negative. When I discussed the issue with the source, they said there is a chance that the cells are contaminated with mycoplasma. We sent some of the cell culture flasks that were more than a week old and they had many of those small particles to the original researchers to test for Mycoplasma and for the second time the results were negative.

I was told the cell number should not exceed 4X10^5 as the cells will kill themselves. I plated fresh vials from liquid Nitrogen into new flasks with different cell number, and the lowest cell concentration, also started showing those small particles/ cells within 24 hours.

I always use a high FBS in my Freeze mix, and when I thaw the cells I add media and spin the cells to eliminate DMSO from freeze mix.

I have worked with human PBMC, monocytes, different cell lines, mouse and rabbit spleen cells, and my cells have always been happy, my cell culture technique is very good and have never ever experienced any contamination in the last 8-9 years.

Advise from someone who has experience working with EBV cells is much appreciated.

Thanks in advance.