Generation and characterization of a high-affinity chimaeric antibody against hepatitis B surface antigen

Bose, Biplab ; Khanna, Navin ; Acharya, Subrat K. ; Sinha, Subrata (2006) Generation and characterization of a high-affinity chimaeric antibody against hepatitis B surface antigen Biotechnology and Applied Biochemistry, 43 (2). pp. 93-101. ISSN 0885-4513

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Antibody against HBsAg (hepatitis B surface antigen) is advocated for the passive immunotherapy in certain cases of hepatitis B infections. A recombinant monoclonal antibody against HBsAg would offer several advantages over the currently used polyclonal human hepatitis B immunoglobulin. 5S is a mouse monoclonal antibody that binds to HBsAg with very high affinity. However, this mouse antibody cannot be used for therapeutic purposes, as it may elicit antimouse immune responses. Chimaerization, by replacing mouse constant domains with human counterparts, can reduce the immunogenicity of this molecule. We have cloned the VH (heavy-chain variable region) and VL (light-chain variable region) genes of this mouse antibody, and fused them with CH1 (heavy-chain constant domain 1) of human IgG1 and CL (light-chain constant domain) of human kappa chain respectively. These chimaeric genes were cloned into a mammalian expression vector (pFab-CMV), which has a modular cassette coding for part of the hinge, CH2 and CH3 of human IgG1. The recombinant construct was transfected in CHO (Chinese-hamster ovary) cells to generate a stable transfectoma. The resulting transfectoma was maintained in a serum-free medium and the full-length chimaeric anti-HBsAg antibody was purified from the culture supernatant. The yield of the purified chimaeric antibody was moderate (≈5.5 mg/l). We further characterized the chimaeric antibody using several in vitro techniques. It was observed that the chimaeric molecule was glycosylated and expressed in the expected heterodimeric form. This chimaeric antibody has very high affinity and specificity, similar to that of the original mouse monoclonal antibody.

Item Type:Article
Source:Copyright of this article belongs to Portland Press.
Keywords:Chimaeric Antibody; Chinese-hamster Ovary Cells (CHO Cells); Hepatitis B Surface Antigen; Monoclonal Antibody; Recombinant; Serum-free Medium
ID Code:62603
Deposited On:22 Sep 2011 02:56
Last Modified:03 Jul 2012 07:52

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