Growth of Chinese hamster ovary (CHO) cells expressing the 5-HT2 serotonin receptor in suspension culture: an efficient method for large-scale acquisition of membrane protein for drug evaluation.
Chinese hamster ovary (CHO) cells expressing the rat 5-HT2 serotonin receptor were grown and evaluated in suspension culture to provide an efficient method of producing membrane-bound receptors for drug discovery. Expression of the 5-HT2 receptor in cells grown in batch suspension culture fluctuated as a function of culture age. Both receptor expression and receptor G-protein coupling were the highest early (1.9 pmol/mg membrane protein) but declined rapidly as the culture increased in age. However, addition of fresh serum-containing medium to stationary-phase cells reversed the decline and, after 24 h of growth, resulted in maximal receptor density and G-protein coupling. This serum response was found to be reproducible and lead to the establishment of a semi-continuous batch culture system in which cells were maintained in a growth state that supported high levels of membrane-incorporated and G protein-coupled receptors. In this system, 50% of the culture volume could be removed at 24-h intervals for membrane preparation and the lost volume replenished with fresh medium, resulting in a continuous supply of high-quality membrane preparations.
Zaworski
P G
PG
Upjohn Company, Kalamazoo, MI 49007, USA.
Evans
D L
DL
Lahti
R A
RA
Gill
G S
GS
D000818:Animals; D001665:Binding Sites; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D005260:Female; D019204:GTP-Binding Proteins; D006160:Guanosine Triphosphate / Q000494:pharmacology; D008565:Membrane Proteins; D010053:Ovary / Q000254:growth & development*; D011985:Receptors, Serotonin / Q000235:genetics*
D008565:Membrane Proteins; D011985:Receptors, Serotonin; D006160:Guanosine Triphosphate; D019204:GTP-Binding Proteins