Fig. 1.

CYP2C22 antibody specificity and liver-specific expression of the enzyme. Polyclonal antibodies were raised to a peptide containing amino acids 366–190 of CYP2C22. (A) Western blots of various hepatic fractions incubated with a 1:3000 dilution of anti-CYP2C22 serum 96 or a 1:3000 dilution of antiserum 97. Before the addition to the blot, the diluted antisera were incubated overnight at 4°C with the indicated amounts of antigenic peptide adsorbed to nitrocellulose paper. Lane 1, total cell extract from primary rat hepatocytes (34 μg); lane 2, rat liver S9 fraction (40 μg); lanes 3 and 4, rat liver microsomes (5 and 15 μg, respectively). (B) Western blots using CYP2C22 antisera 96 (1:1000 dilution) or 97 (1:3000 dilution) or anti-CYP2B1 (1:5000 dilution) against insect cell microsomes expressing rat P450 enzymes (1.5-μg protein), as indicated. (C) Western blot using CYP2C22 antiserum 97 (1:3000 dilution) against microsomes from various tissues isolated from a 230-g male Sprague-Dawley rat. This blot is representative of three blots from different animals. (D) CYP2C22 and 2C11 mRNA expression in the same tissues described in (C), measured by reverse-transcriptase quantitative PCR. Relative mRNA expression is expressed as a percentage of the liver values that were arbitrarily set at 100. Values are means ± S.E.M. of values from three different animals.