A RADIORECEPTOR BINDING ASSAY FOR PLATELET-ACTIVATING-FACTOR (PAF) USING MEMBRANES FROM CHO CELLS EXPRESSING HUMAN PAF RECEPTOR

被引：24

作者：

AOKI, Y ^{[1
]}

NAKAMURA, M ^{[1
]}

KODAMA, H ^{[1
]}

MATSUMOTO, T ^{[1
]}

SHIMIZU, T ^{[1
]}

NOMA, M ^{[1
]}

机构：

[1] UNIV TOKYO, FAC MED, DEPT BIOCHEM, BUNKYO KU, TOKYO 113, JAPAN

来源：

JOURNAL OF IMMUNOLOGICAL METHODS | 1995年 / 186卷 / 02期

关键词：

PLATELET-ACTIVATING FACTOR; RADIORECEPTOR ASSAY; RECOMBINANT HUMAN PLATELET-ACTIVATING FACTOR RECEPTOR;

D O I：

10.1016/0022-1759(95)00147-3

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

A simple and reproducible radioreceptor assay (RRA) has been developed using membranes from CHO cells which can stably express human platelet-activating factor (PAF) receptor. The CHO cells expressing the PAF receptor, termed CHO . 1F8, showed a significant intracellular Ca2+ response to PAF, and the same binding properties to [H-3]WEB 2086, a PAF antagonist, as reported (K-d, 13.6 +/- 1.9 nM; B-max, 2.5 +/- 0.4 pmol/mg protein (n = 6)). A competitive binding assay was done using the CHO . 1F8 cell membranes and [H-3]WEB 2086. The minimum detectable dose of PAF was 0.3 nM (approximate to 30 pg per well) and the assay was highly specific for PAF. This method makes it possible to handle large numbers of samples rapidly and simultaneously, since the receptor membrane is prepared in advance and the binding assay can be completed within 3 h. Using this method, we have determined the production and cell association of PAF in human neutrophils.

引用

页码：225 / 231

页数：7

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