"incomplete antibody" production

a.s.thompson at qub.ac.uk a.s.thompson at qub.ac.uk
Wed Jul 31 09:23:46 EST 2002


I have a problem concerning "complete" versus "incomplete" antibody production from mouse (NS0) hybridomas. Firstly, my background is as a post-doc microbiologist, not immunologist, so apologies if the questions seem naive.

We had a problem with low titer of antibody from cell lines, so I went back and recloned by limiting dilution. Single clones were screened for IgG production by dot blot/alkaline phosphatase, then screened by immunofluorescence (our source of antigen is very limited). Positives were then bulked up to 100 ml volumes; at the 25ml stage, they were checked again for IgG, and all seemed well. I now send these cell lines to a Biotech company who will purify the antibodies, and develop them into a dipstick type assay for us (we're a University Environmental Organisation). The report I get back from them is that in 4 out of the 5 cell lines, most of the IgG present is "incomplete", which they cannot purify. Maybe 5% of the total is "complete". Its suggested its due to the medium used, which was RPMI-1640 (with glutamine), pen/strep, pyruvate, 20% myoclonal FCS. Initially, at first, IL-6 (T-24 superantant) was used as a growth supplement, then eliminated as the cells were bulked up.

Now, why would the cells produce "incomplete" antibody; is it due to the medium used, and if so, can simply switching to a different medium impove the yield of complete antibody? Or, is it a case that I need to go back to the original fusion and start cloning all over again?

Thanks in advance

Andy Thompson
Queens University Belfast.


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