Characterization of carp thrombocytes with specific monoclonal antibodies

Abstract

Monoclonal antibodies (WCL6) specific for carp Cyprinus carpio thrombocytes were produced by immunizing mice with membrane lysates of IgM‐negative peripheral blood leucocytes (PBL) and selected on the negative reaction with B cells. WCL6 was reactive with a membrane molecule of approximately 90 kDa and to a lesser extent with molecules of approximately 95 and 110 kDa. In general, between 30 and 40% of PBL were WCL6 + and appeared to be round to spindle‐shaped cells. Immunohistochemical analysis of cryo‐sections showed much higher numbers of WCL6 + cells in the spleen than in the pronephros, intestine and thymus. Flow cytometric analysis of cell suspensions isolated from these organs only revealed WCL6 + cells (4–10%) in the spleen. Electron microscopy of immunogold‐stained WCL6 + PBL showed round but also some spindle‐shaped cells with canalicular and granular structures, and a more irregular and electron‐dense nucleus than found in lymphocytes. WCL6 + cells with electrondense pyknotic nuclei (without a clear nuclear envelope) were found also and their frequency increased with the length of the isolation and staining procedure used. In the spleen, several differentiation steps of WCL6 + cells were found and hence the spleen seems to be the thrombopoietic organ in carp. Thrombocytes from blood could be activated with collagen; the collagen‐activated cells showed a higher side (90°) scatter by flow cytometric analysis and finally considerable cell death.