Preparation and Application of Monoclonal Antibody Against Human von Willebrand Factor Propeptide.
10.19746/j.cnki.issn.1009-2137.2023.04.042
- Author:
Jiao-Jiao YUAN
1
;
Jie YIN
1
;
Jing LING
2
;
Fei SHEN
1
;
Li-Qian XIE
1
;
Xia BAI
1
;
Chang-Geng RUAN
1
;
Yang HE
3
;
Zhen-Ni MA
4
Author Information
1. Jiangsu Institute of Hematology, National Clinical Research Center for Hematologic Diseases, NHC Key Laboratory of Thrombosis and Hemostasis, The First Affiliated Hospital of Soochow University, Collaborative Innovation Center of Hematology, Soochow University，Suzhou, 215000, Jiangsu Province, China.
2. Department of Hematology and Oncology, Children's Hospital of Soochow University, Suzhou 215000, Jiangsu Province, China.
3. Jiangsu Institute of Hematology, National Clinical Research Center for Hematologic Diseases, NHC Key Laboratory of Thrombosis and Hemostasis, The First Affiliated Hospital of Soochow University, Collaborative Innovation Center of Hematology, Soochow University，Suzhou, 215000, Jiangsu Province, China.Ministry of Education Engineering Center of Hematological Disease, Soochow University, Suzhou 215006, Jiangsu Province, China.
4. Jiangsu Institute of Hematology, National Clinical Research Center for Hematologic Diseases, NHC Key Laboratory of Thrombosis and Hemostasis, The First Affiliated Hospital of Soochow University, Collaborative Innovation Center of Hematology, Soochow University，Suzhou, 215000, Jiangsu Province, China.E-mail: windfr@163.com.
- Publication Type:Journal Article
- Keywords:
VWFpp;
double-antibody sandwich ELISA;
von Willebrand factor
- MeSH:
Animals;
Mice;
Humans;
von Willebrand Factor;
Antibodies, Monoclonal;
Protein Precursors/metabolism*;
von Willebrand Diseases/diagnosis*;
Prognosis
- From:
Journal of Experimental Hematology
2023;31(4):1205-1210
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVE:To develop monoclonal antibodies that can specifically recognize human von Willebrand factor (VWF) propeptide (VWFpp) in plasma, and establish a rapid and reliable method for the detection of VWFpp antigen in plasma by using the double-antibody sandwich ELISA with the obtained anti-VWFpp monoclonal antibody.
METHODS:The recombinant human VWFpp (D1 and D2 regions) protein expressed in eukaryotic cells was used as immunogen to immunize BALB/c mice with routine method, so as to obtain clones of fusion cells. After screening and identification, hybridoma cell lines secreting monoclonal antibodies against VWFpp were selected, and then double-antibody sandwich ELISA assay was used to construct VWFpp antigen detection kit for the determination of VWFpp in human plasma. The levels of VWFpp antigen in plasma of 12 leukemia patients who underwent bone marrow transplantation were dynamically detected.
RESULTS:Two hybridoma cell lines that can be subcultured continuously and secrete monoclonal antibodies against VWFpp were obtained and named SZ175 and SZ176 respectively. Identified by ELISA and Western blot, the antibodies could both specifically recognize VWFpp but couldn't recognize mature VWF (without propeptide). Based on the principle of double-antibody sandwich ELISA, monoclonal antibodies SZ175 and SZ176 were successfully made into a kit for detecting VWFpp antigen. The plasma VWFpp levels of leukemia patients before and after bone marrow transplantation were dynamically detected. The results showed that the plasma VWFpp levels of the patients after transplantation were significantly higher than those before transplantation.
CONCLUSION:Two monoclonal antibodies against VWFpp were successfully prepared, and a double-antibody sandwich ELISA detection kit for VWFpp antigen was constructed, which provides a powerful tool for further study on the biological function of VWFpp, the clinical diagnosis and classification of von Willebrand disease (VWD), and the prognostic monitoring of endothelial injury-related diseases.
