EMIT® Cyclosporine Assay: Development of an application protocol for Technicon AXON® system

Abstract

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 μg/L. Interassay coefficients of variation (CV) were 11.2% ( n = 16; x ̄ = 81 μ g/L ) and 9.4% ( n = 16; x ̄ = 418 μ g/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 μg/L CsA, respectively. Serial dilutions of two patient specimens demonstrated a linear relationship between expected and actual CsA concentrations ( r = 0.996, 0998; regression lines; y = 0.989 x + 11.7; y = 0.979 x + 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® = 1.113 x − 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® = 1.114 x − 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. The EMIT® CsA Assay applied to the Technicon AXON® represents the first application of this assay to a high volume, random access analyzer.