title: [Domestication of suspension CHO cells and its application in the expression of anti-PSMA antibody].
abstract: OBJECTIVE
  To domesticate adherent Chinese hamster ovary (CHO) cells into suspension CHO cells (CHO-S) cultured in serum-free medium,and evaluate the application of the CHO-S cells in antibody expression.
  
  
  METHODS
  Adherent CHO cells were domesticated into CHO-S cells through suspension culture and gradually decreasing the serum concentration, and eventually the cells were cultured in serum-free medium. Based on the anti-prostate-specific membrane antigen ( PSMA) single chain antibody fragment ( Sc Fv) gene sequence obtained from a phage display library, the genes of heavy and light chains were designed, synthesized and cloned into the pc DNA3. 1 vector,and the products were named pc DNA3. 1-HC and pc DNA3. 1-LC respectively. The plasmids were transiently transfected at the ratio of light and heavy chain 3 ∶ 1 into CHO-S cells using Free Style MAX transfection reagent, and the supernatants were harvested at day 7 after transfection. SDS-PAGE and Western blotting were used to detect the antibody expression,and flow cytometry was applied to evaluate its binding activity to PSMA positive cells.
  
  
  RESULTS
  The adherent CHO cells were successfully domesticated into CHO-S cells. Expression plasmids for anti-PSMA antibody heavy chain and light chain were successfully constructed,and anti-PSMA antibody could be secretively expressed in CHO-S cells. Flow cytometry showed that the expressed antibody could specifically bind to PSMA positive cells.
  
  
  CONCLUSIONS
  CHO-S cel s were successful y domesticated from adherent CHO cells, and could be used for antibody expression. This study provided a useful tool for further antibody expression, purification and function study.
authors.forename: Jieheng
authors.initials: J
authors.lastname: Wu
authors.forename: Donghui
authors.initials: D
authors.lastname: Han
authors.forename: Ming
authors.initials: M
authors.lastname: Wei
authors.forename: Guoxu
authors.initials: G
authors.lastname: Zheng
authors.forename: Dian
authors.initials: D
authors.lastname: Jiao
authors.forename: Wenjin
authors.initials: W
authors.lastname: Xi
authors.forename: Angang
authors.initials: A
authors.lastname: Yang
authors.forename: Weijun
authors.initials: W
authors.lastname: Qin
authors.forename: Weihong
authors.initials: W
authors.lastname: Wen
mesh_terms: D000818:Animals; D000906:Antibodies / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism; D000918:Antibody Specificity / Q000276:immunology; D000954:Antigens, Surface / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D015153:Blotting, Western; D016466:CHO Cells; D018929:Cell Culture Techniques; D045744:Cell Line, Tumor; D006224:Cricetinae; D003412:Cricetulus; D005434:Flow Cytometry; D043425:Glutamate Carboxypeptidase II / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D007147:Immunoglobulin Light Chains / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D008297:Male; D011471:Prostatic Neoplasms / Q000276:immunology / Q000378:metabolism / Q000473:pathology; D011485:Protein Binding / Q000276:immunology; D057127:Single-Chain Antibodies / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D014162:Transfection
chemical_list: D000906:Antibodies; D000954:Antigens, Surface; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D057127:Single-Chain Antibodies; C473848:FOLH1 protein, human; D043425:Glutamate Carboxypeptidase II