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This document presents the results of a study comparing the accuracy of a cortisol immunoassay to LC-MS/MS. The study found a significant positive bias in the cortisol immunoassay compared to LC-MS/MS, but could not identify the cause of this difference. However, the study suggests that despite the bias, the cortisol immunoassay may still be considered for adrenal vein sampling (AVS) due to the minimal impact on clinical decision making. The study recommends careful consideration of the positive bias for other indications. The document concludes with acknowledgments to the individuals who contributed to the analysis of the samples. [Extracted from the article]

In clinical practice, steroid measurements are performed mainly by direct, non-extraction immunoassays adapted to high throughput, automated immunoassay platforms and employing secondary calibrators. The accuracy of such steroid immunoassays is limited by cross-reactivity with structurally related steroids and nonspecific matrix interference as well as the metrological traceability of manufacturer supplied calibrators. The accuracy of steroid immunoassay calibrators has been little investigated by independent chemical methods. Steroid concentrations of 41 calibrators (4–6 replicates per calibrator) supplied by four manufacturers for use in testosterone (T), estradiol (E2), and progesterone (P4) commercial immunoassays were measured by ultra-pressure liquid chromatography-mass spectrometry (UPLC-MS). Among 14 non-zero T calibrators, six (43 %) deviated significantly from the label concentration with 29 % outside 20 % of it. Among 14 E2 calibrators, eight (57 %) deviated significantly, whereas seven (50 %) were outside 20 % of the label concentration. Among 11 P4 calibrators, eight (73 %) deviated significantly whereas four (36 %) were outside within 20 % of the label concentration. We conclude that inaccurate calibration of manufacturer's supplied standards may contribute to inaccuracy of commercial direct steroid immunoassays. [ABSTRACT FROM AUTHOR]

Glaucoma is the second most common cause of blindness. An increased intraocular pressure is the only treatable symptom of glaucoma. Because patients often exhibit a poor therapy adherence, a drug depot consisting of ELA-NCO and hyaluronic acid with timolol was developed to ensure sustained drug release. This drug depot is formed by in situ polymerisation after injection into the subconjunctival space. To test the in vivo drug release of timolol in serum and aqueous humour, a liquid chromatography mass spectrometry (LCMS) method was developed and tested using spike- and recovery experiments, and on in vivo samples after topical application. Samples of serum and aqueous humour were taken from New Zealand White rabbits. For topical application, a commercially available formulation of timolol was used. This study presents results concerning the recovery of timolol from spiked samples. Serum and aqueous humour samples were spiked with timolol maleate to a final concentration of 50 ng/mL. Subsequently, the samples were extracted and analysed by LCMS. External calibration of the developed method showed high linearity. Recovery experiments showed no loss of timolol. Hence, the extraction method is robust and able to recover the whole amount of timolol from aqueous humour and serum.

Background: Choosing which biomarker tests to select for further research and development is not only a matter of diagnostic accuracy, but also of the clinical and monetary benefits downstream. Early health economic modeling provides tools to assess the potential effects of biomarker innovation and support decision-making. Methods: We applied early health economic modeling to the case of diagnosing primary aldosteronism in patients with resistant hypertension. We simulated a cohort of patients using a Markov cohort state-transition model. Using the headroom method, we compared the currently used aldosterone-to-renin ratio to a hypothetical new test with perfect diagnostic properties to determine the headroom based on quality-adjusted life-years (QALYs) and costs, followed by threshold analyses to determine the minimal diagnostic accuracy for a cost-effective product. Results: Our model indicated that a perfect diagnostic test would yield 0.027 QALYs and increase costs by €43 per patient. At a cost-effectiveness threshold of €20,000 per QALY, the maximum price for this perfect test to be cost-effective is €498 (95% confidence interval [CI]: €275–€808). The value of the perfect test was most strongly influenced by the sensitivity of the current biomarker test. Threshold analysis showed the novel test needs a sensitivity of at least 0.9 and a specificity of at least 0.7 to be cost-effective. Conclusions: Our model-based approach evaluated the added value of a clinical biomarker innovation, prior to extensive investment in development, clinical studies and implementation. We conclude that early health economic modeling can be a valuable tool when prioritizing biomarker innovations in the laboratory. [ABSTRACT FROM AUTHOR]

Background: Monitoring of plasma drug levels is mandatory in patients receiving high-dose methotrexate. This study evaluated the analytical performance of the novel Architect and the established ARK™ methotrexate immunoassay (running on the Roche Cobas© c502 analyzer) in comparison with liquid chromatography-mass spectrometry (LC-MS) and the TDx/TDxFLx Methotrexate II assay. Methods: Imprecision and linearity were verified for the Architect and ARK assay according to CLSI EP15-A3 and EP6-A guidelines, respectively. The reported limit of quantitation (0.04 μmol/L) was tested for both assays according to the CLSI EP17-A2 guideline. Correlation and agreement between the different assays were evaluated using residual plasma samples (n=153). Results: Total imprecision was <6.3% and <9.5% for the Architect and ARK assay, respectively. The claimed linearity and limit of quantitation were confirmed for the Architect assay. For the ARK assay, imprecision at the limit of quantitation was <18% with a positive bias resulting in a high total error up to 58%, and hence the linearity could not be confirmed. Both assays showed strong correlations with the TDX assay and LC-MS but a positive bias of 12.2% and 20.5% in comparison to LC-MS for the Architect and ARK assay, respectively. For the ARK assay this bias increased dramatically for samples with concentrations towards the limit of quantitation. Conclusions: The Architect assay is suitable for monitoring plasma methotrexate, but the ARK assay showed unsatisfactory performance in the analysis of low concentrated samples. Unlike the TDX assay, both assays require manual dilution of samples at higher concentrations, which delays sample processing in clinical routine. [ABSTRACT FROM AUTHOR]

Background: Accurate cholesterol measurements are essential for the prevention and management of cardiovascular diseases. Quality assurance of cholesterol measurements requires reference methods. Methods: An isotope dilution liquid chromatography mass spectrometric (ID/LC/MS) method was developed. Serum samples were sampled volumetrically using automated dilutors, treated with potassium hydroxide and equilibrated with 3,4-13C2 cholesterol. The natural cholesterol and the internal standard were extracted with hexane and oxidized to cholest-4-en-3,6-diones with chromic acid. The oxidation products were separated on reversed phase LC and detected by tandem MS. The method was calibrated using aqueous cholesterol calibrators and the calibration function was established with a polynomial regression. Results: The correlation coefficients of the calibration curves were always >0.9999. The coefficients of variation (CV) of the volumetric sampling and the LC/MS analysis averaged 0.22% and 0.50%, respectively, and the total measurement CV was 0.60%. Other sources of measurement uncertainty were minor. Results on certified reference materials agreed within 1% of the certified values. Conclusions: An ID/LC/MS method for serum cholesterol has been developed. The method is simple and accurate and may be used as a candidate reference method. [ABSTRACT FROM AUTHOR]