Use of peroxidase:anti-peroxidase immune complexes as markers for Fc gamma receptors in vitro.
In species for which monoclonal antibodies are not yet available, the demonstration of Fc receptors has relied on a variety of ligand-based assays including the binding of antibody-coated erythrocytes, radiolabeled monomeric immunoglobulin, and non-physiologic aggregates of immunoglobulin. In order to study the binding of small immune complexes to Fc receptors on canine monocytes, a new method was developed using an enzyme-linked immune complex. The ability of rabbit polyclonal peroxidase:anti-peroxidase (PAP) immune complexes to bind to freshly isolated canine peripheral blood monocytes was characterized using standard ELISA techniques. The binding of rabbit polyclonal PAP to monocytes was time and concentration dependent and reversible. This binding was saturable with increasing concentrations of PAP and could be blocked by soluble rabbit IgG or rabbit Fc fragments. The blocking and saturation curves for canine monocytes were suggestive of multiple classes of Fc gamma binding sites. In contrast to intact PAP complexes, the binding of F(ab) PAP preparations or free horseradish peroxidase was minimal. The use of commercially available PAP preparations provides a reproducible, inexpensive, and non-radioactive measure of Fc gamma receptor binding on canine cells. In addition, these findings suggest caution in using heterologous PAP as a histochemical reagent in tissues expressing Fc gamma receptors.
Kreisle
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Department of Veterinary Pathobiology, Purdue University, West Lafayette, IN 47907-1243, USA.
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D000818:Animals; D000936:Antigen-Antibody Complex / Q000276:immunology*; D001667:Binding, Competitive / Q000276:immunology; D015415:Biomarkers; D004285:Dogs / Q000097:blood*; D004797:Enzyme-Linked Immunosorbent Assay / Q000662:veterinary; D007124:Immunoenzyme Techniques / Q000662:veterinary*; D007128:Immunoglobulin Fragments / Q000276:immunology; D007074:Immunoglobulin G / Q000276:immunology; D009000:Monocytes / Q000276:immunology*; D011817:Rabbits; D017452:Receptors, IgG / Q000276:immunology*
D000936:Antigen-Antibody Complex; D015415:Biomarkers; D007128:Immunoglobulin Fragments; D007074:Immunoglobulin G; D017452:Receptors, IgG