title: A comparative study of polyclonal and monoclonal antibodies for immunocytochemical localization of cytosolic aspartate aminotransferase in rat liver.
abstract: The rabbit antiserum and mouse monoclonal hybridoma antibody against porcine cytosolic aspartate aminotransferase (c-AAT) (or cytosolic glutamic oxaloacetic transaminase (c-GOT)) were produced and compared for the localization of c-AAT in rat liver. An indirect immunocytochemical technique was performed using peroxidase-conjugated goat immunoglobulin (Ig) G anti-rabbit IgG and peroxidase-conjugated rabbit IgG anti-mouse IgG as the second antibody. Rats were perfused with paraformaldehyde-lysine-periodate fixative and the liver fragments were immersed in 4% paraformaldehyde and transferred to 10% dimethyl sulfoxide overnight and subjected to cryostat sectioning. The rabbit IgG antibody, 3 individual monoclonal antibodies, and a mixture of these 3 monoclonal antibodies were applied to the tissue sections, respectively, using the same concentration. Under the same experimental conditions, the c-AAT was localized in each individual hepatocyte by both monoclonal and polyclonal antibodies. However, a mixture of three monoclonal antibodies gave stronger staining than a single monoclonal antibody; although two antibodies yield more intense staining than just one, it was still less intense than for three. The conventional rabbit polyclonal antibody against c-AAT produced more reaction product than the combined three monoclonal antibodies. It is concluded that for immunocytochemical study, the use of a single monoclonal antibody is sensitive enough to localize its tissue antigen under the present experimental condition. To obtain a stronger reaction product, a combination of several monoclonal antibodies, at least three or more, may give better staining.
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000032:analysis / Q000276:immunology*; D000918:Antibody Specificity; D001219:Aspartate Aminotransferases / Q000032:analysis / Q000276:immunology*; D003600:Cytosol / Q000201:enzymology* / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D007106:Immune Sera / Q000032:analysis / Q000276:immunology; D008099:Liver / Q000201:enzymology* / Q000276:immunology; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D011817:Rabbits; D051381:Rats; D011919:Rats, Inbred Strains; D013552:Swine
chemical_list: D000911:Antibodies, Monoclonal; D007106:Immune Sera; D001219:Aspartate Aminotransferases