Is there a more stringent elution buffer than 2% SDS with BME? I have tried this both at RT and 50 degrees with very little antibody eluted on SDS-Page. Based on ELISA, the antibodies are definitely still bound to the antigen-coupled beads.
Eluted antibodies do not need to be functional, but should be able to be visualized on SDS-Page.
If you have the antibody bound to a column, you could try electroelution of the protein from the gel. This method is used for affinity-purification of immunoglobulins, to avoid getting mediocre antibodies (poor antibodies do not bind to the column, high affinity ones are not eluted). It also avoids the use of protein denaturants.
Engelbert Buxbaum Thanks for answering; the beads are magnetic so electroelution may not be an option. I also would like to capture all antibodies (including poor/high affinity ones) as I am analyzing an antigen-specific IgG repertoire.
Boiling the beads in SDS-PAGE sample buffer should elute everything. If not, try including 6M urea in the sample buffer. Since reducing conditions are used, each IgG will appear as 2 bands (heavy and light chain), but since there are many different antibodies in an antiserum, all slightly different and with varying degrees of glycosylation, you will actually see broad bands centered at about 50 and 25 kDa.
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