Growing hybridoma cells in medium without serum - (Jul/06/2011 )

Dear all,

I would like to raise a question regarding the growth of hybridoma cells in media without serum. So far, I have been using the classical RPMI + 10% SFB for the culture of these cells. However, since I learned my antibodies of interest get contaminated with undesired bovine immunoglobulins, I decided to try to adapt my hybridomas to these new fashionable media .
I have just started with the process and my idea is to mix the new medium with gradually decreasing proportions of the classical one (according to the manufacturer´s specifications). However, something that remains unclear is whether once I succeed in getting my cells to grow in the new medium, I should replace the medium daily.
I split my cells every 2-3 days with RPMI+serum. Can I do the same with the serum-free medium if I keep the same cell densities?
Please, note that what I want to know is whether the medium has to be changed just because of its unstability, regardless cell density.
I would greatly appreciate any comments on this.
Thanks in advance!

My guess is that it depends on the medium. We use the serum free media exactly the same as media with added serum. The cells do however grow more slowly in the serum free medium.

Hello Chelo,

The short answer to your question is that you may continue to passage your cells every 2-3 days according to cell density until your cells are used to the serum free media formulation. Here is my long answer ---- As you already know, because of the interference of the contaminating bovine antibodies, your antibodies need to be produced in a serum free system. And going serum free means getting your cells used to a completely different environment, in the absence of serum.And I think you are already in the process of adapting your cells by passaging in this new serum free media formulation.However, this adaptation process depends on the serum free media formulation that you choose. If your cells are not accustomed to the formulation, it may take more than 2-3 days for the cells to grow to enough density to warrant a cell passage. Sometimes, they even lose viability in that you have to culture more cells to recover from such a drop in cell viability.

I helped develop in the lab, an animal free, completely recombinant and contaminanting IgG (and any other Ig contamination) free FBS replacement supplement called ZAP-Hybridoma. This supplement is very easy to use because you can use it with an optimal basic classical media (suitable for serum free formulation)without a lengthy adaptation stage. I currently work with hybridoma groups gloablly who have successfully implemented this supplement into their hybridoma processes. They have reported short periods of adaptation (as little as a direct switch to the serum free formulation without a substantial drop in cell viability). You can use this supplement without sacrificing the performance that you are accustomed to with your serum, and at the same time avoid the bovine contamination. Lastly, when we designed this product, we made it cost effective. So, you will save money as well.

Please feel free to contact me at msantos@invitria.com if you would like to discuss this further. I sincerely hope that you will find this supplement useful in your goal to convert to a serum free system. Thank you very much.