Enhanced specificity due to method specific limits for relative ion intensities in a high-performance liquid chromatography – tandem mass spectrometry method for iohexol in human serum

• Published in: Clinical chemistry and laboratory medicine Vol. 58; no. 5; pp. 709 - 718
• Main Authors: Schweinsberg, Vincent, Eckelt, Stefan, Schulz, Anna Marta, Arlt, Marcel, Schumann, Marc, van der Giet, Markus, Tauber, Rudolf, Binscheck-Domaß, Torsten, Kießling, Peggy
• Format: Journal Article
• Language: English
• Published: Germany De Gruyter 01.05.2020; Walter De Gruyter & Company
• Subjects: Acceptance criteria; Chemical Precipitation; Chromatography; Chromatography, High Pressure Liquid - methods; Chromatography, High Pressure Liquid - standards; Computer simulation; Elution; Glomerular Filtration Rate; High performance liquid chromatography; Humans; iohexol; Iohexol - analysis; Iohexol - isolation & purification; Iohexol - standards; Ions; kidney function; Kidney Function Tests - methods; Limit of Detection; Liquid chromatography; Mass spectrometry; Mass spectroscopy; Mathematical analysis; Matrix methods; Reference Standards; relative ion intensities; Reproducibility of Results; Scientific imaging; Selectivity; Sensitivity enhancement; Serum - chemistry; Spectroscopy; tandem mass spectrometry; Tandem Mass Spectrometry - methods; Tandem Mass Spectrometry - standards; Training; high-performance liquid chromatography; iohexol; kidney function; tandem mass spectrometry; glomerular filtration rate; relative ion intensities
• ISSN: 1434-6621; 1437-4331
• DOI: 10.1515/cclm-2019-1001

Background Accurate assessment of kidney function is needed for a variety of clinical indications and for research. The measurement of the serum clearance of iohexol has emerged as a feasible method to reach this objective. We report the analytical validation and clinical application of a new high-performance liquid chromatography (HPLC) - tandem mass spectrometry (MS/MS) assay to quantify iohexol in human serum. Specificity was enhanced due to the use of method specific acceptance limits for relative ion (RI) intensities. Methods The internal standard ioversol was added to 50 μL serum prior to protein precipitation with methanol. Linear gradient elution was performed on a Waters Oasis® HLB column. Three transitions for both iohexol and ioversol were monitored allowing calculation of RIs. Measurements acquired during method validation were used as a training set to establish stricter acceptance criteria for RIs which were then tested retrospectively on clinical routine measurements (86 measurements) and on mathematically simulated interferences. Results The method was linear between 5.0 μg/mL (lower limit of quantification [LLOQ]) and 100.3 μg/mL iohexol. Intraday and interday imprecision were ≤2.6% and ≤3.2%, respectively. Bias was -1.6% to 1.5%. All validation criteria were met, including selectivity, recovery, extraction efficiency and matrix effects. Retrospectively acceptance limits for RIs could be narrowed to ±4 relative standard deviations of the corresponding RIs in the training set. The new limits resulted in an enhanced sensitivity for the simulated interferences. Conclusions Criteria for validation were met and the assay is now used in our clinical routine diagnostics and in research.