Antibody from hen's eggs against a conserved sequence of the gametophytic self-incompatibility proteins of plants

A convenient way of producing effective antibodies to plant proteins is presented. It takes into account the following facts: (1) many plant proteins are highly glycosylated, thus giving rise to nonspecific antibodies which cross-react with other glycoproteins, (2) it is more common nowadays to know the DNA sequence of a protein coding gene than to have significant amounts of that protein well purified as antigen, and (3) eggs from immunized hens are very convenient sources of large amounts of antibodies. An antibody which specifically detects self-incompatibility proteins (S-proteins) in Solanaceae was isolated. The published cDNA sequences and deduced amino acid sequences of Nicotiana alata (an ornamental tobacco) S-proteins were computer analyzed in order to recognize a peptide with a conserved sequence which would effectively give rise to antibodies. An adequate amount of this peptide was synthesized and part of it was coupled to a macromolecular carrier (bovine serum albumin). Antibodies to this peptide-carrier complex were recovered from egg yolks of immunized hen in amounts corresponding to 300 ml of antiserum per month. From the total immunoglobulin fraction isolated, the antibody specific for S-proteins was affinity purified over 100-fold on a peptide-coupled column. The antibody showed high levels of specificity for putative S-specific sequences in N. alata and in petunia. Also the size, polymorphy, and quantity of the detected antigens were in very good accordance with previously published data. In addition to the present application, this procedure should be useful for a wide variety of proteins.