EMIT cyclosporine assay: development of an application protocol for Technicon AXON System

Monitoring parent drug cyclosporine (CsA) concentrations in whole blood has been facilitated by the introduction of automated nonisotopic immunoassays [fluorescence polarization monoclonal whole blood assay (FPIA), EMIT Cyclosporine Assay]. The latter assay currently has a defined application only for Cobas Mira Chemistry Systems. The purpose of our work was to develop an application for this assay on the Technicon AXON. Instrument settings were optimized to arrive at the following assay performance characteristics. Limit of sensitivity was 50 micrograms/L. Interassay coefficients of variation (CV) were 11.2% (n = 16; mean = 81 micrograms/L) and 9.4% (n = 16; mean = 418 micrograms/L). Recoveries of 102, 112, and 117% were obtained by spiking aliquots of 10 whole blood patient pools of known CsA concentrations with 50, 100, and 200 micrograms/L CsA, respectively. Serial dilutions of two patient specimens demonstrated a linear relationship between expected and actual CsA concentrations (r = 0.996, 0.998; regression lines; y = 0.989x + 11.7; y = 0.979x + 9.5). Carryover and interference (lipemia) were not evident. Instrument calibration stability is at least 1 month. Comparison with CsA concentrations analyzed in renal transplant patients by the FPIA assay produced a linear regression equation of EMIT = 1.113x - 44.5, r = 0.968, Sy/x = 20.8, n = 32. Comparison with high-performance liquid chromatography (HPLC)-derived values in the same patient population produced a linear regression equation of EMIT = 1.114x - 16.4, r = 0.970, Sy/x = 20.2. FPIA-derived CsA concentrations averaged 14.2% more than those obtained with the EMIT method with the latter averaging 1.3% more than HPLC values.(ABSTRACT TRUNCATED AT 250 WORDS)