Isolation and purification of bovine IgM by dissociating immunoglobulin-brucella complexes.
Methods for the dissociation of immunoglobulin-Brucella complexes (IgM-rich) were evaluated. The antigen-antibody complex was separated from serum by centrifugation and dissociated with either H2O, 2.0 M pH 2.0 glycine buffer, or various concentrations of KSCN. Optimal antibody recoveries for KSCN were obtained using 1.0--2.0 M KSCN. Dissociation of the complex with water resulted in the highest recovery of antibody and the glycine buffer was lowest. Immunoelectrophoretic analysis of the eluted antibody revealed the presence of IgG and IgM. At concentrations of KSCN greater than 2.5 M, albumin-like protein was also eluted from the antibody-antigen complex. The IgM could readily be separated from IgG and albumin by gel filtration. Purified IgM was Brucella specific, monodisperse, and had a sedimentation coefficient of 20.0.
D000818:Animals; D000936:Antigen-Antibody Complex; D000942:Antigens, Bacterial; D002003:Brucella abortus / Q000276:immunology*; D002417:Cattle; D002850:Chromatography, Gel; D005998:Glycine / Q000494:pharmacology; D006041:Goats; D005779:Immunodiffusion; D007122:Immunoelectrophoresis; D007075:Immunoglobulin M / Q000302:isolation & purification*; D007136:Immunoglobulins; D007158:Immunologic Techniques; D011817:Rabbits; D013861:Thiocyanates / Q000494:pharmacology; D013997:Time Factors
D000936:Antigen-Antibody Complex; D000942:Antigens, Bacterial; D007075:Immunoglobulin M; D007136:Immunoglobulins; D013861:Thiocyanates; D005998:Glycine