[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.

CONCLUSION

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.