TY - JOUR T1 - Development of an AntiPeptide Antibody That Binds to the C-Terminal Region of Human CYP1B1 JF - Drug Metabolism and Disposition JO - Drug Metab Dispos SP - 274 LP - 280 VL - 27 IS - 2 AU - Yong Ming Tang AU - Gen-Fu Chen AU - Patricia A. Thompson AU - Dong-Xin Lin AU - Nicholas P. Lang AU - Fred F. Kadlubar Y1 - 1999/02/01 UR - http://dmd.aspetjournals.org/content/27/2/274.abstract N2 - An antipeptide antibody was raised against a 14-mer synthetic peptide (CDFRANPNEPA KMN) corresponding to the amino acid sequence from 491 to 504 of human cytochrome P-450 (CYP)1B1. Rabbit-derived antisera demonstrated the ability to induce moderately high antibody titers (>1:105) as judged by enzyme-linked immunosorbent assay. In Western blot analysis, the purified antibody recognized a single protein band (estimated as 56 kDa) in microsomes prepared from human and rodent tissues. No significant cross-reactivity to either human CYP1A1 or human CYP1A2 protein was detected. Titration studies using recombinant human CYP1B1 and an enhanced chemiluminescence-based detection method demonstrated a minimal detection sensitivity for this antiserum at about 0.34 ng/band in 8 × 7-cm minigels. The immunoprecipitation and immunoinhibition results indicate that this antisera recognizes the nondenatured human CYP1B1 protein but does not inhibit its enzyme activity. Using this antibody, CYP1B1 protein was detected in nine different human tissues and in cultured cells induced by various chemicals. This highly specific, highly sensitive antibody provides an important tool to study tissue distribution and cellular expression levels of CYP1B1, with negligible cross-reactivity from the other members of the CYP1 family. The American Society for Pharmacology and Experimental Therapeutics ER -