PURIFICATION AND CHARACTERIZATION OF HIGH-MOLECULAR-WEIGHT (HMW) AND LOW-MOLECULAR-WEIGHT (LMW) IMMUNOGLOBULINS FROM THE SERUM OF THE DOGFISH, SCYLIORHINUS-CANICULA, AND THE IDENTIFICATION OF ANTIBODY-FORMING TISSUES

1. High molecular weight (HMW) and low molecular weight (LMW)-like Ig was purified and characterized from dogfish, Scyliorhinus canicula serum. 2. HMW Ig was isolated by gel filtration on a Sepharose 6B column and by fractionation on an anion exchange Mono Q FPLC column. The LMW-Ig was separated on a rabbit anti-dogfish Ig immunoaffinity column and a Zorbax G-250 HPLC gel filtration column. 3. Both LMW and HMW-Ig forms had identical heavy and light chains of 70 kDa and 25 kDa respectively on reducing SDS-PAGE. Intact HMW-Ig had a Mr similar to pentameric human IgM of 952 kDa, while the LMW-Ig was 160-200 kDa as estimated by non-reducing SDS-PAGE. 4. Anti-keyhole limpet haemocyanin serum was raised in dogfish and fractionated on a gel filtration Sepharose 6B column. The immunoreactivity fractions corresponded to the HMW-Ig peak with a Mr of ca 950 kDa. 5. Specific antiserum was raised against dogfish HMW-Ig in rabbits and employed to identify antibody secreting cells in the spleen, Leydig and epigonal organs of the adult dogfish.