When using polyclonal antibodies to detect an array of host cell proteins what changes to that polyclonal antibody require a recharacterization. More specifically would the following situation warrant a recharacterization: if the non specific binding of the polyclonal antibody is high and, in attempts to lower the background, the antibody is run over a column (that does not contain that antigens of interest).
(10-10-2015, 09:44 PM)Deneb Wrote: [ -> ]When using polyclonal antibodies to detect an array of host cell proteins what changes to that polyclonal antibody require a recharacterization. More specifically would the following situation warrant a recharacterization: if the non specific binding of the polyclonal antibody is high and, in attempts to lower the background, the antibody is run over a column (that does not contain that antigens of interest).
Hi Deneb,
Should I assume that your main goal is to get rid of the HCPs from your targeted therapeutic product? If it is so, then I hope the polyclonal antibodies were developed using the null cell of the same host? If you did so, your polyclonal antibodies should be specific enough for HCPs. Further purity can be induced by taking care of the cross-reactivity (if your therapeutic product is a MAb especially).
Your proposed situation for recharacterization does seem interesting as it might saturate most of the non-specific receptors/ antibodies. Worth trying but I its utility would depend upon your ultimate goal.
Can you share more info about the source of your pAbs/ type of product you are aiming at? anything that you think might help..
Sunil,
Your assumption is right that the goal is to remove HCP from a therapeutic product. This is a generic assay used for many different products. The pAbs are derived from goats immunized with null cell lines. My concern is that when running the pAbs over a column to reduce background is if I see a reduction in high end binding. This would make me think I am losing a portion of the specific antibodies as well. For this reason I would be compelled to ensure that my coverage has remained the same (compare coverage pre-adsorbtion and post-adsorption).
Thank you for your input.