Evaluation of a novel semi-automated HPLC procedure for whole blood cyclosporin A confirms equivalence to adjusted monoclonal values from Abbott TDx.
The problem in the measurement of cyclosporin (CyA) is that the widely used immuno-based assays suffer from interference by metabolites present in unpredictable excess. To resolve this, the consensus view has been to develop more specific and robust procedures for the measurement of CyA alone in order to give values similar to those obtained by HPLC. We developed an alternative strategy based on Abbott poly- and monoclonal assays to derive an adjusted monoclonal value as an equivalent measurement to HPLC. We have now evaluated a recently developed semi-automated HPLC procedure and used it to test the validity of the adjusted monoclonal value. The automated HPLC procedure with online clean-up was optimised for the separation of CyA and internal standard CyD. The assay was simple to use, precise and gave good recovery of cyclosporin from whole blood. Comparisons with the more specific immunoassays Abbott AxSym and EMIT showed close agreement, whereas Abbott monoclonal values indicated up to 20% positive bias. In contrast, the adjusted monoclonal values gave good agreement with HPLC. Data obtained from HPLC linked to tandem mass spectrometry (MS) indicated closer agreement with Abbott monoclonal values than expected, suggesting some positive bias with MS. The benefit of using an adjusted monoclonal value is that a result equivalent to HPLC is obtained, as well as an indication of the concentration of metabolites from the Abbott polyclonal measurement.
Roberts
Norman B
NB
Department of Clinical Biochemistry, The Royal Liverpool and Broadgreen University Hospitals, Liverpool, UK. n.b.roberts@liverpool.ac.uk
Dutton
John
J
Higgins
Gerald
G
Allars
Lesley
L
D000906:Antibodies; D000911:Antibodies, Monoclonal; D001331:Automation; D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation / Q000379:methods*; D016572:Cyclosporine / Q000097:blood* / Q000378:metabolism; D003524:Cyclosporins / Q000592:standards; D008401:Gas Chromatography-Mass Spectrometry; D007118:Immunoassay; D007166:Immunosuppressive Agents / Q000276:immunology / Q000378:metabolism; D011933:Reagent Kits, Diagnostic; D012015:Reference Standards; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization
D000906:Antibodies; D000911:Antibodies, Monoclonal; D003524:Cyclosporins; D007166:Immunosuppressive Agents; D011933:Reagent Kits, Diagnostic; D016572:Cyclosporine; C050026:cyclosporin D