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BackgroundRheumatoid arthritis (RA) is a systemic autoimmune disease affecting approximately 1% of the population [1]. The pathogenesis of RA involves the overexpression of tumor necrosis factor alpha (TNFα) and other cytokines [2]. Adalimumab (ADA) is a human monoclonal antibody directed against TNFα and is highly effective in the treatment of RA. For efficient treatment trough levels of ADA need to be adjusted within a therapeutic window which is 5 to 10 μg/mL [3]. A rapid test allows faster reporting of trough levels, providing a great advantage over test formats that need samples to be sent to a central or service laboratory. Here we report on the technical performance evaluation of the Quantum Blue® Adalimumab lateral flow test.ObjectivesDevelopment and performance evaluation of a rapid test for the monitoring of ADA trough levels in human serum at the point of care.MethodsThe sandwich lateral flow immunoassay uses a TNFα coated gold label and a highly specific monoclonal antibody immobilized on the test membrane to detect ADA in diluted human serum samples. Sensitivity of the assay was determined by calculating limit of detection (LoD) and limit of quantification (LoQ) according to CLSI EP17-A2 guideline. Moreover, the assay was evaluated regarding cross-reactivity with other therapeutic antibodies targeting TNFα, influence of rheumatoid factors (RF) and high dose hook effect. A method comparison was performed against a commercially available ELISA (RIDASCREEN® ADM Monitoring, R-Biopharm, Germany) to compare the trough level results of 40 patients treated with ADA. All statistical analyses were performed with Analyse-it for Excel.ResultsThe Quantum Blue® Adalimumab test allowed analysis of serum samples within 15 minutes. The samples were diluted 1:20 in chase buffer before application onto a test cassette (volume 80 μL). The readout was performed with the Quantum Blue® Reader resulting in adalimumab concentration levels in the lower μg/mL range. The test exhibited a LoD of 0.2 μg/mL and a LoQ of 0.69 μg/mL. No high dose hook effect was detected for samples containing up to 1000 μg/mL ADA. The latter two data sets allowed a measuring range of 1 to 35 μg/mL of ADA in patient samples. Other therapeutic TNFα blockers, like infliximab and golimumab, showed no cross-reactivity with the Quantum Blue® Adalimumab test. RF showed no influence on correct measurement of ADA at all tested concentrations. The method comparison to a well-established commercial ELISA method revealed a slope of 1.12 and a regression coefficient (r2) of 0.90 (by Passing-Bablok). A Bland-Altman analysis showed a bias of 1.9% confirming the overall excellent correlation of the two methods as well as the accuracy of our newly developed rapid test.ConclusionsThe BÜHLMANN Quantum Blue® Adalimumab assay enables the quantitative determination of ADA trough levels over the clinically relevant range in serum with a time to result of only 15 minutes. The assay exhibits an excellent accuracy and correlation to a well-established laboratory reference me...

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