[Expression of human-mouse chimeric antibody against CD40 in CHO cell line and characterization of its function].
OBJECTIVE
To investigate the stable expression of a chimeric antibody against CD40 moleculeèch-5C11éin CHO and its biological activity.


METHODS
Human-mouse chimeric antibody against CD40 recombinant plasmid and mock plasmid were transfected into CHO cell line through lipofectamine mediation. Human kappa chain and Fc fragment of ch-5C11 were characterized by FACS and Western blot. The concentration of ch-5C11 in cell supernatants was detected by Lowry assay and the inhibitory effect of ch-5C11 on the proliferation of Daudi cells was detected by MTT assay.


RESULTS
RT-PCR showed that target CHO cells integrated chimeric heavy chain and chimeric light chain gene. FACS and Western blot showed that ch-5C11 in cell supernatants maintained the binding activity and specificity to human CD40 molecule, and contained human kappa chain and Fc fragment. Cell supernatants were purified using protein G affinity chromatography. The concentration of human-mouse chimeric antibody against CD40 in cell supernatants was 0.535 mg/L. When co-cultured with B lymphoma cell line Daudi, ch-5C11 induced proliferation arrest of Daudi cells.


CONCLUSIONS
The human-mouse chimeric antibody against CD40 can be expressed in CHO stably and effectively, which inhibits proliferation of Daudi.
Qu
Qiu-xia
QX
Key Laboratory of Clinical Immunology of Jiangsu Province, First Affiliated Hospital, Soochow University, Suzhou 215006, China.
Chen
Cheng
C
Wang
Qin
Q
Ge
Yan
Y
Chen
Yong-jing
YJ
Qiu
Yu-hua
YH
Zhang
Xue-guang
XG
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology*; D019013:CD40 Antigens / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D016466:CHO Cells; D002478:Cells, Cultured; D006224:Cricetinae; D003412:Cricetulus; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D051379:Mice; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
D000911:Antibodies, Monoclonal; D019013:CD40 Antigens; D007141:Immunoglobulin Fc Fragments; D011993:Recombinant Fusion Proteins