Ying Gu1, Shun-dong Ji, Yi-ming Zhao, Fei Shen, Chang-geng Ruan. 1. Jiangsu Institute of Hematology, The First Affiliated Hospital of Soochow University, Key Laboratory of Thrombosis and Hemostasis, Ministry of Health, Suzhou 215006, China.
Abstract
AIM: To prepare and identify a monoclonal antibody (mAb) against human platelet glycoprotein Ib and make its application. METHODS: BALB/c mice were immunized with human platelets washed, and the spleen cells of them were fused with myloma cells. A hybridoma cell was screened by indirect ELISA and cloned, and the mAb were purified from the ascites of mice. Ig subclass was analysed by double immunodiffusion. The antigen recognized by monoclonal antibody was identified by flow cytometry and radioimmunoassy, respectively. The inhibition of mAb on plasma von Willebrand factor ristocetin cofactor activity (vWF:Rcof) was investigated by enzyme linked immunosorbent assay (ELISA). RESULTS: A murine mAb against human platelet membrane glycoprotein (GP) Ib was developed and denominated as SZ-151. SZ-151 belonged to IgG1 subclass and its titer in ascites was 1:20 000. Flow cytometry and radioimmunoassy showed that the antigen recognized by monoclonal antibody SZ-151 was platelet membrane GPIb. ELISA showed that SZ-151 did not inhibit plasma von Willebrand factor ristocetin cofactor activity. CONCLUSION: A mAb, SZ-151 against platelet glycoprotein Ib was developed, which could be useful in assays of plasma von Willebrand factor ristocetin cofactor activity(vWF:Rcof) and can be used for diagnose patients with vWD.
AIM: To prepare and identify a monoclonal antibody (mAb) against human platelet glycoprotein Ib and make its application. METHODS: BALB/c mice were immunized with human platelets washed, and the spleen cells of them were fused with myloma cells. A hybridoma cell was screened by indirect ELISA and cloned, and the mAb were purified from the ascites of mice. Ig subclass was analysed by double immunodiffusion. The antigen recognized by monoclonal antibody was identified by flow cytometry and radioimmunoassy, respectively. The inhibition of mAb on plasma von Willebrand factor ristocetin cofactor activity (vWF:Rcof) was investigated by enzyme linked immunosorbent assay (ELISA). RESULTS: A murine mAb against human platelet membrane glycoprotein (GP) Ib was developed and denominated as SZ-151. SZ-151 belonged to IgG1 subclass and its titer in ascites was 1:20 000. Flow cytometry and radioimmunoassy showed that the antigen recognized by monoclonal antibody SZ-151 was platelet membrane GPIb. ELISA showed that SZ-151 did not inhibit plasma von Willebrand factor ristocetin cofactor activity. CONCLUSION: A mAb, SZ-151 against platelet glycoprotein Ib was developed, which could be useful in assays of plasma von Willebrand factor ristocetin cofactor activity(vWF:Rcof) and can be used for diagnose patients with vWD.