The TDx assay for cyclosporine and its metabolites in blood samples compared with HPLC and RIA methods.

Two methodologies have been developed to monitor cyclosporine (CsA) therapy: high-performance liquid chromatography (HPLC) and radioimmunoassay (RIA). Recently, a fluorescence polarization immunoassay (FPIA) has also become commercially available for the assay of CsA and its metabolites. The authors compared the results obtained with a modified FPIA with those found with two RIAs which use a polyclonal antibody, in order to verify if the FPIA assay is suitable for routine measurements in blood samples. Moreover, the accuracy of the RIAs and of the modified FPIA was checked against the results obtained by an HPLC technique assumed as a reference assay. The FPIA assay for CsA in blood samples seems preferable to the RIAs; in fact, as far as specificity is concerned, the TDx assay is comparable to polyclonal RIAs, while the precision (both within- and between-laboratories) is significantly better. Moreover, the TDx method is easier and faster to perform (20 samples can be assayed in about 30 min, while 2-4 h are necessary with RIA), with fewer handling steps; the instrumentation is automated and the reagents are more stable and less hazardous than those used in RIA.

Evaluation and comparison of radioimmunoassay methods using monoclonal or polyclonal antibodies for the assay of cyclosporine in blood samples.

Results obtained measuring blood Cyclosporine A (CsA) concentrations in transplanted patients (124 samples of cardiac, 20 samples of liver, and 10 samples of kidney transplanted patients) by the use of two monoclonal radioimmunoassay (RIA) methods have been compared with those found using the HPLC technique (considered as the reference method) and two polyclonal RIAs. In addition, results on quality control samples collected in a multicentre collaborative study for CsA assay from the users of the same monoclonal and polyclonal RIAs were analysed to evaluate the performance of the methods under study. Polyclonal RIAs, which measure both the parent molecule and its metabolites, produced results 1.5-3 times higher than HPLC or monoclonal RIAs. On the contrary the two RIAs, which use monoclonal antibodies specific for CsA, show a better correlation with HPLC; these RIAs, which measure the intact drug molecule only, are recommended when the monitoring of the native molecule of CsA is requested. As far as the reproducibility is concerned, the four RIAs (both polyclonal and monoclonal) exhibit an unsatisfactory degree of between-assay and between-lab precision, since the coefficients of variation (CVs) ranged from 19.4% to 23.1%.