The properties of mabs are more diverse than thought like other proteins. Things like glycosylation and pI can make huge differences in the traditional Protein A or G purification.

Have you tried both Protein A and G?

Have you tried basic elution rather than acidic?

Thanks for reply, I use protein A for rabbit host and G for mouse. No I haven't tried different one for same antibody. Elution buffer in my kit (GEHC) is acidic, don't know how to make basic buffer. How basic helps? Thanks 

Your protein / mAb may be precipitating out at the wrong pH 

I would tray a mix protein A/G for simplicity can also do salt elutions 

Thanks, How do I know which PH is the right one for specific antibody?

Is the low yield due to denaturation of the antibodies? Would probably lead to precipitates in the tubes. One alternative then could be to add 1M Tris buffer to the elution tubes prior to the elution to rapidly neutralize the eluted antibodies. 1 M Tris pH 7,5 can neutralize several volumes of eluted antibodies.

If you use antigen specific affinity purification, the column may bleed blocking part of the antibodies. Look for  antigen in the eluate. In that case you will need primary labelled antibodies for the detection of the antigen.

Not stringent enough conditions (antibodies remain on the column). Change to high pH as suggested above or elute at pH 2,5.

Another possibility is of course that there are low amounts of antibodies in the starting material. 

There was an old booklet made by Pharmacia called affinity purification techniques (blue on the front if I remember correctly). You may have some old copies at GE Healthcare in Uppsala if you look around.

Hi, Thanks for the feedback. The specific antibody that I'm struggling, is Mouse Monoclonal antibody IgG1, in liquid tissue culture supernatant containing sodium azide. Total protein 3.7g/L, and Ig 261mg/L in concentrate form according to vendor. When I purify with protein G HT, It seems I loose all through washes, because I don't get any after with elutions. I'm not chemist and don't have that much experience. My Binding buffer is 0.2M sodium phosphate,PH7.0, Elution buffer is 1M glycine-HCI,PH2.7, and I add 1M Tris-HCI,PH9 to elution tubes, all provided in a kit. Wash tubes has lots of protein, but elution nothing. I appreciate all the help I can get. I tried diluting the washes with binding buffer and running through the column again, results the same.

I think you should either change to protein G sepharose or increase the pH for the application and washing. Monoclonal mouse IgG1 is notorius for not binding to protein A, even if polyclonal mouse IgG normally always binds to protein A at neutral pH.

If I remember correctly the article below had recommendations for protein A purification of mouse IgG monoclonals. Unfortunatelly, I do not have the full article.

Lindmark R, Thorén-Tolling K, Sjöquist J. Binding of immunoglobulins to protein A and immunoglobulin levels in mammalian sera. J Immunol Methods. 1983 Aug 12;62(1):1-13.

You could have a look at this protein A handbook that recommends binding at pH 8-9.

See link below

https://www.abdserotec.com/static/marketing/proteinahandbook2-205524.pdf