中国仓鼠卵巢细胞
抗体
分子生物学
生物
免疫球蛋白轻链
单克隆抗体
人源化抗体
重组DNA
细胞培养
病毒学
免疫学
基因
遗传学
作者
Daniel A. Mueller,Lars Heinig,Sanja Ramljak,Astrid Krueger,Reiner Schulte,Arne Wrede,Andreas W. Stuke
出处
期刊:Hybridoma
[Mary Ann Liebert]
日期:2010-12-01
卷期号:29 (6): 463-472
被引量:2
标识
DOI:10.1089/hyb.2010.0041
摘要
Because of their high antigen specificity and metabolic stability, genetically engineered human monoclonal antibodies are on the way to becoming one of the most promising medical diagnostics and therapeutics. In order to establish an in vitro system capable of producing such biosimilar antibodies, we used human constant chain sequences to design the novel human antibody expressing vector cassette pMAB-ABX. A bidirectional tetracycline (tet)-controllable promotor was used for harmonized expression of immunoglobulin type G (IgG) heavy and light chains. As an example we used anti-prion protein (anti-PrP) IgGs. Therefore, the variable heavy (VH) and light chain (VL) sequences of anti-PrP antibodies, previously generated in our laboratory by DNA immunization of prion protein knock-out mice, were isolated from murine hybridoma cell lines and inserted into pMAB-ABX vector. After transfection of Chinese hamster ovary (CHO) cells, a number of stable antibody producing cell clones were selected. One cell line (pMAB-ABX-13F10/3B5) stably expressing the recombinant humanized antibody (rechuAb) 13F10/3B5 was selected for detailed characterization by Western blot, immunofluorescence, and flow cytometric analyses. The full-length recombinant humanized IgG antibody showed a high level of expression in the cytoplasm. In conclusion, the new cell system described here is a suitable tool to produce functional intact full-length humanized IgG antibodies.
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