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, radiolab eled monomeric immunoglobulin, and non-physiologic aggregates of immun oglobulin. In order to study the binding of small immune complexes to Fe receptors on canine monocytes, a new method was developed using an enzyme-linked immune complex. The ability of rabbit polyclonal peroxid ase:anti-peroxidase (PAP) immune complexes to bind to freshly isolated canine peripheral blood monocytes was characterized using standard EL ISA techniques. The binding of rabbit poly clonal PAP to monocytes was time and concentration dependent and reversible. This binding was sat urable with increasing concentrations of PAP and could be blocked by s oluble 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 reproduc ible, inexpensive, and non-radioactive measure of Fc gamma receptor bi nding on canine cells. In addition, these findings suggest caution in using heterologous PAP as a histochemical reagent in tissues expressin g Fc gamma receptors.