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Concerning Cultivation of Monoclonal Antibodies - How long before antibodies spoil in media? (Sep/01/2010 )

Hello all, I have a question I hope somebody with experience with hybridomas could answer.
I'm trying to produce a large quantity of hybridoma supernatant for future antibody isolation, and I am a little concerned about spoiling of the antibodies.

I've never grown hybridoma cells before (Neither has anyone in the department). I purchased a vial of frozen cells and have been cultivating them up to 2 L of AB laden supernatant. From starting from a 0.5 mL vial of frozen cells, I have split and resplit flasks and eventually combined everything into a 2.5 L Roller bottle.
However, from beginning to finish (Right now the cells are showing approx 50% viability via Trypan Blue), it has been a total of about 4-5 weeks. I am wondering if this has been too long and if my antibodies could possibly be spoiled by now, and what I could do in the future to speed up growth of cells (If I need to reculture)?

-David Cochran-

Hi David, this seems a normal time length to me. I used to collect supernatant regularly in separate sterile culture flask, storing them at 4 Degree Celsius. When having collected enough supernatant I would then directly proceed to purifying and concentrating the antibody. For re-culturing just make sure you use cells that were frozen after low passage number previously. Good luck!

-rana pequena-