Instituto Interuniversitario de Investigación de Reconocimiento Molecular y Desarrollo Tecnológico (IDM), Universitat Politècnica de València-Universitat de València, 46022 Valencia, Spain.
Departamento de Microbiología, Facultad de Medicina, Universidad de València, Av. Blasco Ibáñez 17, 46010 València, Spain.
Anal Chem. 2023 Aug 15;95(32):12113-12121. doi: 10.1021/acs.analchem.3c02284. Epub 2023 Aug 6.
The global prevalence of β-lactam allergy poses a major challenge in administering first-line antibiotics, such as penicillins, to a significant portion of the population. The lack of β-lactam IgE antibody pools with defined selectivity hampers the standardization and validation of in vitro assays for β-lactam allergy testing. To address this limitation, this study introduces a synthetic IgE specific to β-lactam antibiotics as a valuable tool for drug allergy research and diagnostic tests. Using phage display technology, we constructed a library of human single-chain antibody fragments (scFv) to target the primary determinant of amoxicillin, a widely used β-lactam antibiotic. Subsequently, we produced a complete human synthetic IgE molecule using the highly efficient baculovirus expression vector system. This synthetic IgE molecule served as a standard in an in vitro chemiluminescence immunoassay for β-lactam antibiotic allergy testing. Our results demonstrated a detection limit of 0.05 IU/mL (0.63 pM), excellent specificity (100%), and a four-fold higher clinical sensitivity (73%) compared to the in vitro reference assay when testing a cohort of 150 serum samples. These findings have significant implications for reliable interlaboratory comparison studies, accurate labeling of allergic patients, and combating the global public health threat of antimicrobial resistance. Furthermore, by serving as a valuable trueness control material, the synthetic IgE facilitates the standardization of diagnostic tests for β-lactam allergy and demonstrates the potential of utilizing this synthetic strategy as a promising approach for generating reference materials in drug allergy research and diagnostics.
β-内酰胺类抗生素过敏在全球的流行，对相当一部分人群使用一线抗生素（如青霉素）造成了重大挑战。缺乏具有明确选择性的β-内酰胺 IgE 抗体库，阻碍了用于β-内酰胺类抗生素过敏检测的体外检测方法的标准化和验证。为了解决这一限制，本研究引入了一种针对β-内酰胺类抗生素的合成 IgE，作为药物过敏研究和诊断检测的有价值工具。我们使用噬菌体展示技术，构建了一个针对阿莫西林主要决定簇的人源单链抗体片段（scFv）文库，阿莫西林是一种广泛使用的β-内酰胺类抗生素。随后，我们使用高效的杆状病毒表达载体系统产生了完整的人源合成 IgE 分子。该合成 IgE 分子在用于β-内酰胺类抗生素过敏检测的体外化学发光免疫测定中作为标准品。我们的结果表明，与体外参考检测相比，当检测 150 份血清样本的队列时，该检测方法的检测限为 0.05 IU/mL（0.63 pM），具有出色的特异性（100%）和四倍更高的临床灵敏度（73%）。这些发现对于可靠的实验室间比较研究、过敏患者的准确标记以及对抗全球公共卫生威胁的抗生素耐药性具有重要意义。此外，作为有价值的真值控制材料，该合成 IgE 有助于β-内酰胺类抗生素过敏诊断检测的标准化，并展示了利用这种合成策略作为药物过敏研究和诊断中生成参考材料的有前途方法的潜力。
