Homogenous enzyme immunoassay for cyclosporine in whole blood using the EMIT (R) 2000 cyclosporine specific assay with the COBAS MIRA-Plus analyzer

We describe the evaluation of the EMIT (R) 2000 cyclosporine specific assay kit, an enzyme-multiplied immunoassay for cyclosporine in whole blood, wit h a COBAS MIRA-plus analyzer. The enzyme used for the assay was glucose-6-p hosphate dehydrogenase (EC 1.1.1.49 G6PDH) from Leuconostoc mesenteroides; the monoclonal antibody is fairly specific for cyclosporine, and is not rea ctive with most metabolites. The assay principle is based on competitive im munoassay with G6PDH-labeled cyclosporine and cyclosporine in sample to the anticyclosporine mouse monoclonal antibody binding site. The within-assay coefficient of variation (CV) of this method was 2.7-4.2% (n = 10) at the l evels of 56.2-339.7 mug/L. Day-to-day CVs ranged from 4.2-8.1 % at the leve ls of 47.2-350.2 mug/L. The within-day CVs ranged from 2.0-6.4% at the leve ls of 43.3-330.5 mug/L. The functional detection limit was 24.9 mug/L. Samp les treated with pretreatment reagent were stable at least 5 hr. Calibratio n was stable at least 10 days. The analytical recovery was 81-109%. The cor relation between values obtained with the EMIT 2000 cyclosporine specific a ssay kit (y) and fluorescence polarization immunoassay (FPIA) (TDxFLx) (x) was: y = 0.880x - 13.053 mug/L (r = 0.984, Sy/x = 15.968, n = 71) with a me an difference of 31.42 +/- 19.89 mug/L ((TDxFLx-EMIT (R) 2000) +/- SD); for the FPIA (AxSYM) (x): y = 0.989 - 4.144 mug/L (r = 0.981, Sy/x = 17.478, n = 71) with a mean difference of 5.56 +/- 17.38 mug/L ((AxSYM-EMIT 2000) SD ); and for the radioimmunoassay (RIA, CYCLO-Trac SP) (x): y = 0.893-6.764 m ug/L (r = 0.993, Sy/x =10.582, n = 71) with a mean difference of 22.18 +/- 14.98 mug/L ((RIA- EMIT (R) 2000) +/- SD) using the Bland-Altman technique. J. Clin. Lab. Anal. 15:319-323, 2001. (C) 2001 Wiley-Liss, Inc.