Doud Emma H, Hansen Kasi, Haynes Kathryn A, Eldridge Kierra, Arrivalagan Jaison, Charbe Nitin, Da Silva Lais, Quinney Sara K, Mosley Amber L, Rizzo Stacey J Sukoff, Territo Paul R
School of Medicine, Indiana University, Indianapolis, IN, USA.
School of Medicine, University of Pittsburgh, Pittsburgh, PA, USA.
MAbs. 2025 Dec;17(1):2537118. doi: 10.1080/19420862.2025.2537118. Epub 2025 Jul 22.
A sensitive and specific liquid chromatography-tandem mass spectrometry assay was developed for the quantification of chimeric aducanumab (chAdu), a therapeutic antibody targeting pathological amyloid plaques in Alzheimer's disease, in murine biological matrices. This method addresses the challenges of quantifying biotherapeutics in multiple tissue types within preclinical animal models with complex genetic backgrounds, where traditional enzyme-linked immunosorbent assay (ELISA) methods may suffer from interference and limited sensitivity. The assay uses parallel reaction monitoring on a Lumos Tribrid Orbitrap mass spectrometer, coupled with an Evosep LC system, and AssayMap Bravo-based automated sample processing. Key features include protein A enrichment for improved sensitivity, optimized peptide selection based on sequence uniqueness and ionization response, and incorporation of stable isotope-labeled peptides for accurate quantification. Assay performance was evaluated for selectivity, repeatability, and stability. The fit-for-purpose assay was successfully applied to quantify chAdu in both mouse cortex and plasma samples obtained from a pilot pharmacokinetic study of a mouse model of amyloid plaque deposition. This targeted mass spectrometry workflow offers a robust and reproducible alternative to ELISA for preclinical biotherapeutic analysis, particularly when dealing with complex biological samples.
开发了一种灵敏且特异的液相色谱-串联质谱分析法，用于在小鼠生物基质中定量检测嵌合型阿杜卡单抗（chAdu），这是一种靶向阿尔茨海默病病理性淀粉样斑块的治疗性抗体。该方法解决了在具有复杂遗传背景的临床前动物模型中，对多种组织类型中的生物治疗药物进行定量的挑战，在这种情况下，传统的酶联免疫吸附测定（ELISA）方法可能会受到干扰且灵敏度有限。该分析方法在Lumos Tribrid Orbitrap质谱仪上采用平行反应监测，结合Evosep液相色谱系统和基于AssayMap Bravo的自动化样品处理。关键特性包括利用蛋白A富集提高灵敏度、基于序列独特性和电离响应优化肽段选择，以及掺入稳定同位素标记的肽段进行准确定量。对该分析方法的选择性、重复性和稳定性进行了评估。该适用的分析方法成功应用于定量检测从小鼠淀粉样斑块沉积模型的初步药代动力学研究中获得的小鼠皮质和血浆样品中的chAdu。这种靶向质谱工作流程为临床前生物治疗分析提供了一种强大且可重复的ELISA替代方法，特别是在处理复杂生物样品时。
