Measurement of cyclosporine in plasma from patients with various transplants: HPLC radioimmunoassay with a specific monoclonal antibody compared.

Abstract

This study compares cyclosporin A (CsA) concentrations in plasma from patients receiving various transplants, as measured by HPLC and RIA with a monoclonal antibody for CsA and an 125I-labeled ligand. The RIA was restandardized with in-house standards because it overestimated CsA by an average of 23%. The RIA was sensitive to 2 micrograms/L, the standard curve was linear from 20 to 500 micrograms of CsA per liter, analytical recovery was 98%, and CVs were less than 8% for intra- and interassay precision. RIA (y) vs HPLC (x) for 283 plasma samples from 145 patients gave a slope = 1.1256, r = 0.979. When the results were segregated according to transplant type, CsA in liver and heart recipients was overestimated by RIA as compared with HPLC: slope = 1.202, r = 0.973 and slope = 1.1477, r = 0.983, respectively. Adult and pediatric CsA values were acceptable when RIA and HPLC were compared: slope = 1.0755, r = 0.977 and slope = 1.0563, r = 0.980, respectively. For six samples (four heart, two liver recipients) where HPLC and RIA values demonstrated wide discrepancies, repeat HPLC and analysis of eluate fractions gave CsA concentrations nearer values by the initial HPLC assay. We conclude that this RIA cannot be substituted for HPLC in the case of heart and liver recipients. The need for each laboratory to standardize the RIA is obvious.

Bioequivalence study between two formulations of ciclosporin A (Cyclavance® oral solution and Atopica® soft capsules) following a single oral administration to dogs

“…Furthermore, sampling times chosen in accordance with the EMA guidelines and the use of a previously validated LC-MS/MS method for measuring blood ciclosporin A levels, enabled to accurately characterize the plasma concentration-time profile of the two drugs. Indeed, LC-MS/MS has been described as one of the most sensitive, reliable and fast analytical technique for bioequivalence studies [ 15 , 16 ] and is commonly used for determining ciclosporin A levels [ 1 ]. When Cyclavance® oral solution and Atopica® soft capsules were administered orally at a target dose of 5 mg/kg body weight of ciclosporin A, the parametric 90 % confidence intervals of the mean ratio test/reference were actually included within the reference confidence interval [0.80–1.25] for Cmax and AUC0-t parameters.…”

Drug‐Drug Interaction After Single Oral Doses of the Furanocoumarin Methoxsalen and Cyclosporine

“…Plasma concentrations of cyclosporine were measured by a monoclonal antibody fluorescence polarization immunoassay (mFPIA) using TDx (Abbott Diagnostics, Roodepoort, South Africa) 37 . Plasma was obtained by standard protocol after reequilibrating whole blood to 37°C by incubating for 30 minutes in a water bath, followed by immediate centrifugation 38 . Compared to whole‐blood analyses, determination of plasma cyclosporine concentrations is acceptable and preferred at many centers, with the understanding from clinicians that plasma levels are typically 2.1‐fold lower than whole‐blood levels.…”

“…37 Plasma was obtained by standard protocol after reequilibrating whole blood to 37°C by incubating for 30 minutes in a water bath, followed by immediate centrifugation. 38 Compared to whole-blood analyses, determination of plasma cyclosporine concentrations is acceptable and preferred at many centers, with the understanding from clinicians that plasma levels are typically 2.1-fold lower than whole-blood levels. The use of mFPIA is standard practice for many therapeutic drug monitoring laboratories and provides an acceptably rapid, precise, and accurate means to measure cyclosporine concentrations.…”

A Practical Method for the Synthesis of a Cyclosporine−Fluorescein Conjugate

“…Immunoassays using radiolabeled , or fluorescently labeled − cyclosporine are preferable as analytical methods, due to their rapid assay time and technical ease. These assays have been particularly useful in discriminating between CsA and its generally inactive metabolites due to the employment of selective monoclonal antibodies that show specificity, reproducibility, and sensitivity. , In the case of covalent labeling reactions, synthetic processes for CsA conjugates are lengthy or low yielding, particularly for coupling via the MeBmt residue due to steric factors caused by CsA's three-dimensional conformation. , We report here a straightforward, scaleable process that provides reproducible yields of a CsA−fluorescein conjugate linked through the hydroxy group of the MeBmt.…”