Patent ID: 6063582
Filing Date: 2000-05-16
Classification: A61K,C07K,G01N

Abstract:
A method to determine whether a candidate compound activates or inhibits a P.sub.2U2 receptor, comprising the steps of:providing a host cell transformed or injected with a nucleic acid molecule which encodes a P.sub.2U2 receptor and which hybridizes to the complement of a nucleic acid molecule having SEQ ID NO:1 under conditions selected from the group consisting of: (1) washing with 0.015 M NaCl, 0.0015 M sodium titrate, 0.1% NaDodSO.sub.4 at 50.degree. C.; (2) hybridization in 50% (vol/vol) formamide with 0.1% bovine serum albumin, 0.1% Ficoll, 0.1% polyvinylpyrrolidone, 50 mM sodium phosphate buffer at pH 6.5 with 750 mM NaCl, 75 mM sodium citrate at 42.degree. C.; and (3) hybridization in 50% formamide, 5.times.SSC (0.75 M NaCl, 0.075 M sodium citrate), 50 mM sodium phosphate (pH 6.8), 0.1% sodium pyrophosphate, 5.times.Denhardt's solution, sonicated salmon sperm DNA (50 .mu.g/ml), 0.1% SDS, and 10% dextran sulfate at 42.degree. C., with washes at 42.degree. C. in 0.2.times.SSC and 0.1% SDS, whereby said host cell expresses on its surface a P.sub.2U2 receptor;contacting said host cell with the candidate compound;measuring the compound's effect on the P.sub.2U2 receptor expressed by the host cell;comparing the compound's effect on the P.sub.2U2 receptor to the compound's effect on a control cell; anddetermining whether the compound activates or inhibits the P.sub.2U2 receptor expressed by said encoding nucleic acid molecule, wherein said P.sub.2U2 receptor is activated by four agonists in the following order of specificity: UTP>UDP>ADP>ATP in Xenopus laevis oocytes.