Evaluation of glycoprotein Ib expression on feline platelets

Abstract

Objective - To determine whether platelets obtained from cats expressed glycoprotein Ib (GPIb). Sample Population - Platelets obtained from 11 specific-pathogen-free cats. Procedure - Platelets were analyzed by use of immunofluorescence microscopy, flow cytometry, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, western immunoblot analysis, and immunoprecipitation. Results - Immunofluorescence microscopy and flow cytometry revealed the protein on the surface of feline platelets. Biochemical studies (western immunoblot analysis and immunoprecipitation) revealed a 140-kd membrane glycoprotein. Additional biochemical studies revealed that feline GPIb was sensitive to proteolysis, because platelet cytoskeletons prepared with low concentrations of a calpain inhibitor (ie, leupeptin; 100 μg/ml) had substantial proteolysis, and there was an association of protein fragments with the actin cytoskeleton. Conclusions and Clinical Relevance - Analysis of these results indicate that feline platelets express a 140-kd membrane protein that is recognized by monoclonal antibodies developed against GPIb. Application of standardized ELISA to quantitate glycocalicin, the water-soluble fragment of GPIb, may provide important information on the production of microvesicles, increased platelet turnover, and abnormal proteolysis.