Isolation and Characterization of Somatic and Metabolic Antigens and Moulting Fluid Antigens of 'Dirofilaria immitis' Adults, Microfilaria, Late First Stage (Sausage-Form) and Third Stage Larvae

Dirofilaria immitis adult, cuticular and microfilarial antigens have been obtained by a combination of phosphate buffer, 1% Triton X-100, 12% Triton X-100, butanol extractions and papain digestion. A number of adult, cuticular, and microfilarial antigens have been isolated by affinity chromatography. Antigen protein heterogenicity has been monitored by isoelectric focusing electrophoresis in polyacrylamide gels employing a pH gradient of 3-10. A protein-antigen coated polyacrylamide microbead agglutination test by antibody has been developed. Naturally infected dog sera in some cases in dilutions of up to 1:10,000 have agglutinated protein-antigen coated polyacrylamide microbeads. Absorption of sera antibodies by affinity chromatography serving as controls failed to agglutinate the microspheres. Antigens have been partially characterized by isoelectric focusing electrophoresis in polyacrylamide gels. Electrophoretically separated gels at pH 3-10, pH 3-6, pH 5-8 and pH 7-10 have been sectioned into 2 mm wafers. Eluted proteins were assayed fluorometrically for total protein content and by soluble antigen fluorescent antibody for antigens. (Author)