purification of OKT3

Nathalie Jacobs N.Jacobs at ulg.ac.be
Mon Aug 26 06:31:38 EST 1996

we try to purify the mouse antibody OKT3 (IgG2a) from the hybridoma 
supernatant. We are using a home made sepharose A column (non HPLC). 
We have several problems: 
-a contamination with bovin IgG (the hybridoma is cultivated with RPMI 
1640 + 20% SVF. Have somebody already try to cultivate them with 
hybridoma medium without serum?)
-we don't obtain a lot of antibodies at the pH 4.5 (the pH for IgG2a). 
We obtain higher concentration at pH 5.5(certainly bovin IgG)! We try 
to concentrate the supernatant but everything have been detached at pH 
I would appreciate any suggestion to increase the concentration of 
purified OKT3.
Thank you in advance.
Nathalie Jacobs

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