Preparation and identification of monoclonal antibody against angiopoietin-related protein 2 / 第二军医大学学报

ABSTRACT

Objective:To prepare a monoclonal antibody against human angiopoietin-related protein 2(ARP2).Methods: Human spleen ARP2 gene was obtained by RT-PCR.A pET32a-ARP2 plasmid was constructed and was incorporated into(E.coli.) The products were purified and were used to immunize 6-week-old BALB/c female mice.Hybridoma secreting anti-ARP2 monoclonal antibody was obtained by standard procedure.Mass production was carried out after specificity identification with Western blotting.Results: The fusion protein obtained by pET32a system had a relative molecular weight of about 570 000,which was in accordance with the theoretical value.The purity of the protein was more than 90% after purification.The antibody titer was 110~(4)in the hybridoma culture supernatant and 110~(7)-10~(8) in the ascites.The IgG2a type antibody had a relative molecular weight of about 570 000 by Western blot analysis.Immunohistochemistry method showed that the antibody bond with human ARP2.Conclusion: The prepared anti-human ARP2 monoclonal antibody in this study can be used for identification of ARP2 protein.