Patent ID: 6410340
Filing Date: 2002-06-25
Classification: G01N,Y10S

Abstract:
A protein binding assay method for the immunosuppressant drugs FK-506, rapamycin, and pharmacologically active metabolites or derivatives thereof (â€œanalytesâ€ ), in a fluid sample, comprising the steps of:(a) contacting said fluid sample with an isolated about 8.4 kDa binding protein (â€œimmunophilinâ€ ) so that a complex forms between said analyte and said binding protein; (b) quantifying the amount of analyte specifically bound to said binding protein; and, (c) relating the amount of said analyte specifically bound to the concentration of said drug in said fluid sample; wherein said isolated about 8.4 kDa immunophilin exhibits the following characteristics: (i) the first 23 amino acid sequence is identical to that of authentic ubiquitin; (ii) retention times on HPLC are identical to those of authentic ubiquitin; (iii) migrates on SDS-PAGE plates identically to HPLC-purified commercial ubiquitin; (iv) exhibits a Kd of about 0.8 nM for FK-506 and 0.08 nM for rapamycin; (v) has a Hill plot value of about unity; and (vi) when complexed to FK-506 inhibits calcineurin phosphatase activity.