Re: what is the difference between a monoclonal and polyclonal antibody?

Hi Jimmy-

Ah, what a wonderful question, and part of it is not as simple as it might seem. I'll answer the easy part first - a monoclonal antibody is the result of a single B-cell - cancer-cell fusion, that is grown up into a clonal population (thus mono-clonal). The important thing here is that the monoclonal antibody produced will have identical idiotype/specificity (it is lots of one antibody). Monoclonals are usually produced by immunizing an animal, then removing the spleen and fusing it with a non-immunoglobin- producing B-cell cancer cell, forming a hybridoma. If the hybridoma makes antibody, the antibody-producing genes are from the spleen B cell, not the cancer B cell, but the cells will grow indefinitely in culture due to the transformed nature of the cancer B cell. The hybridomas are then cloned by limiting dilution or FACS-based single cell sorting and screened for desirable antibody production. So that's a monoclonal antibody.

A polyclonal antibody is actually a mix of different antibodies (poly (many) - clonal). Polyclonals are usually generated by immunizing an animal and then removing the "immune serum" and purifying the immunoglobulin-containing fraction. These antibody mixes recognize multiple different epitopes (because they are generated by a whole pool of B cells, rather than a single one).

But you also asked about specificity, and this is the difficult part, because it depends on the antibody. If you have a high-specificity monoclonal, it could be more specific than a polyclonal (and often cleaner in western blot due to the lack of "contaminating" antibodies in polyclonal sera that are not directed against your protein of interest). However, many proteins do not lend themselves to good monoclonal generation, either because they are highly conserved, or otherwise are poorly immunogenic. In this case, a polyclonal can be a better choice, as the aggregate "avidity" of the polyclonal serum is higher than any single antibody. Another case where polyclonals are helpful is if your protein of interest occurs in many forms. In this case, change in protein conformation, phosphorylation, ubiquitinylation, etc., can mask the epitope that the monoclonal antibody binds, and you'll lose reactivity. In this case, you should have "coverage" in a polyclonal, due to the multiple epitopes the polyclonal antibodies recognize.

Hope this helps, and good luck with your westerns...

ILD
MadScientist