Production of human antithrombin-III in a serum-free culture of CHO cells.
A simple method was developed to establish serum-independent Chinese hamster ovary (CHO) cells that grew and secreted high level of human antithrombin-III (AT-III). First, human AT-III and mouse dihydrofolate reductase (DHFR) cDNAs were transfected into DHFR-deficient CHO cells. Transfected cells were treated with increasing concentrations of methotrexate (MTX) and clones secreting high levels of AT-III (10-20 micrograms/ml/3 day) in a serum-containing medium were obtained. Serum-independent clones were derived from the serum-dependent clones by simply culturing the cells for a few weeks in a serum-free medium. In a serum-free medium the established serum-independent clones grew at normal rate and produced almost equivalent amount of AT-III to that of the serum-dependent, parent clones. In addition, AT-III from the serum-independent clones has specific activity similar to that of plasma-derived AT-III.
Yamauchi
T
T
Biotechnology Group, Green Cross Corporation, Osaka, Japan.
Asakura
E
E
Amatsuji
Y
Y
Uno
S
S
Furuta
R
R
Tujikawa
M
M
Tanabe
T
T
D000818:Animals; D000990:Antithrombin III / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D016466:CHO Cells / Q000378:metabolism*; D006224:Cricetinae; D016895:Culture Media, Serum-Free; D004247:DNA / Q000096:biosynthesis; D005822:Genetic Vectors; D006801:Humans; D008099:Liver; D051379:Mice; D010957:Plasmids; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification
D016895:Culture Media, Serum-Free; D011994:Recombinant Proteins; D000990:Antithrombin III; D004247:DNA