Multiplexed analytical approaches to beta-lactam allergy in vitro testing standardization

The suspicion of beta-lactam allergy directly contributes to the prescription of antibiotics that diverge from the guidelines, increasing antimicrobial resistance, one of the biggest threats to global health. In vitro quantification of specific IgE is very useful for monitoring allergy, as it confirms or rules out immediate beta-lactam drug allergy and helps find safe alternative antibiotic stewardship. However, reliable in vitro quantification of specific IgE to beta-lactam antibiotics by immunoassay is challenging because of the difficulty of having selective immunoreagents, mainly beta-lactam antigens, and its low concentration levels in serum. Thus, reliable and sensitive in vitro tests for multiplex detection of allergy to different beta-lactam antibiotics is currently essential for clinical diagnosis. Nevertheless, the lack of standardization of quantitative in vitro methods makes the comparison and interpretation of the results difficult. Here, as proof of concept, we report an improved multiplex microimmunoassay for beta-lactam allergy in vitro testing standardization. The results revealed that homologous calibration allows reliable quantification of specific IgE in human serum at very low concentrations (144 ng L−1). Moreover, the reproducibility of the results increases 2-fold using an internal standard, achieving accurate quantitative information: 93% and 106% recovery for penicillin and amoxicillin, respectively. We simultaneously evaluated the reliability of the improved multiplexed in vitro method in a cohort of 40 human serum samples and achieved excellent agreement (0.99) with a currently used in vitro test. [Display omitted] •The impact of the homologous calibration is demonstrated by testing specific IgE to β-lactam antibiotics.•Internal homologous calibration provides inter-method comparison with 0.99 agreement for penicillin G and amoxicillin.•A chimeric IgE is used as internal standard to improve the analytical performance of the β-lactam in vitro testing.•A sensitivity of 144 ng/L, 2-fold better reproducibility and 93-106% of accuracy is reached.•A reliable medical care at the point-of-need to expand delabeling initiatives in a cost-effective way is achieved