[A method for improving the accuracy and sensitivity of cell membrane chromatography].
OBJECTIVE
To improve the accuracy and sensitivity of cell membrane chromatography (CMC) and evaluate the feasibility of CMC in the study of subtype receptors.


METHODS
Plasmids were used to transfer alpha(1B)-AR cDNA into human embryonic kidney 293 (HEK293) cell lines to obtain cell lines stably overexpressing the subtype receptors. HEK293 alpha(1B) cell membrane stationary phase (CMSP) was prepared by immobilizing the cell membrane on silica. The retention time of 9 alpha(1)-adrenoceptor ligands and capacity factors(kappa'(HEK293 alpha1B)) were calculated. The capacity factors of rat liver tissue and primary cultured rat hepatocytes were also calculated for a correlation analysis.


RESULTS
The calculated capacity factors (kappa') were positively correlated to the published pKi values. The affinity rank orders were identical. The longest retention of the 9 alpha(1)-adrenoceptor ligands occurred on CMSP prepared with HEK293 alpha(1B) cell lines, while CMSP obtained from rat liver tissue showed the shortest retention of the ligands.


CONCLUSIONS
CMC proves practical in the study of the subtype adrenoceptors. The accuracy and sensitivity of CMC can be improved using HEK293 alpha(1B) cell membrane.
Wang
Yu
Y
Xi'an Jiaotong University College of Medicine, Xi'an 710061, China.
Wang
Yu
Y
Deng
Xiu-ling
XL
Yuan
Bing-xiang
BX
He
Lang-chong
LC
D000818:Animals; D002462:Cell Membrane / Q000378:metabolism*; D002846:Chromatography, Affinity / Q000379:methods*; D018076:DNA, Complementary / Q000378:metabolism; D005260:Female; D057809:HEK293 Cells; D006801:Humans; D007668:Kidney / Q000166:cytology / Q000196:embryology; D008024:Ligands; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley; D018340:Receptors, Adrenergic, alpha-1 / Q000378:metabolism*; D012680:Sensitivity and Specificity
C547716:ADRA1B protein, human; D018076:DNA, Complementary; D008024:Ligands; D018340:Receptors, Adrenergic, alpha-1