This forum seems to concentrate on HPLC, but hopefully the experts can help with this problem.
We are having problems eluting our protein of interest from an immunoaffinity column prepared in house. Running buffer is 50mM Tris pH7 and elution buffer 0.1M Citrate pH3, the column is a Hitrap NHS-activated sepharose linked with our protein A-purified antibody. The column appears to work for a few runs but then loses capacity altogether. We have tried different washes, including 0.2M Acetic acid, 2M NaCl, and 3M Thiocyanate, with no effect. When the resin was boiled in sample buffer and ran on SDS/PAGE we see our protein of interest. Clearly the protein is sticking very strongly to the antibody (the same material is applied to another column with a different antibody with no loss of capacity).
My question is what elution conditions should we try? Would McIlvaines buffer at pH2.5/2.6 work better, or should we go with a lower/higher pH? Are we likely to damage the antibody and/or our target protein if we use these extreme pHs? Is there a regeneration step that might help? Finally, would there be any benefit in using other components in the elution buffer, such as salt or detergent?

Looks like the blind have to help the lame again. We have done affinity chrom. only as part of kits, so it is not known what the elution buffer contains. However, I am surprised that you used citrate as that is quite lyotropic. My guess is that one should use something that solvates the protein, namely a chaotropic substance, like your thiocyanate. Apparently you used this thiocyanate, etc., after you had citrate on the column?
Normally the antibody antigen interaction amounts to only a few kcals, they should come off unless something drastic happened. The proteins with which we have worked stayed natured only at pH near 7+ and ionic strength of ~300mmol.
There are some substances sold for renaturing proteins, my suggestion: check Pierce, google.com, and cyclodextrins.
It would certainly be instructive to be informed about the solution to your problem.