de Souza Israel Donizeti, Fernandes Grecco Caroline, Costa Queiroz Maria Eugênia, Pichon Valerie, Combès Audrey
Department of Analytical, Bioanalytical Sciences and Miniaturization (LSABM), Chemistry, Biology and Innovation (CBI), UMR 8231 ESPCI Paris - CNRS, ESPCI Paris, PSL University, 10 Rue Vauquelin, 75005, Paris, France.
Departamento de Química da Faculdade de Filosofia, Ciências E Letras de Ribeirão Preto (DQ-FFCLRP), Universidade de São Paulo, Ribeirão Preto, São Paulo, Brazil.
Anal Bioanal Chem. 2025 Sep 1. doi: 10.1007/s00216-025-06085-7.
The ratio between beta-amyloid (Aβ) peptides 40 and 42 is recognized as a biomarker for Alzheimer's disease, playing a significant role in early diagnosis and disease progression monitoring. Aβ peptides are present at trace levels in cerebrospinal fluid, therefore, developing a new selective extraction procedure is essential for isolating targeted biomarkers from the matrix interferents, ensuring accurate identification and quantification. In this study, a hybrid organic-silica monolith was synthesized in a 530 µm inner diameter-capillary and used for the covalent grafting of beta amyloid peptide aptamers. The resulting miniaturized oligosorbent (mOS) was applied to selectively extract Aβ peptides 40 and 42 from artificial cerebrospinal fluid (CSF) samples. The immobilization procedure achieved grafting yields higher than 90% leading to a dense coverage of Aβ aptamers (655 + 15 pmol.µL of oligosorbent, n = 3, RSD = 2.3), and in a capacity exceeding 50 pmol.µL of mOS. After optimization in pure media, an extraction recovery of 74% and 31% for Aβ40 and Aβ42 peptides, respectively was reached on this mOS. The developed method including extraction on mOS and LC-MS analysis achieved LLOQ values of 0.1 ng.mL, precision and accuracy with CV and RSD values ranging from 1.0% to 12.9% and -4.7% to 11.1%, respectively. This method was successfully applied to selectively extract Aβ peptides from artificial CSF samples, effectively isolating the two Aβ targeted peptides from this complex biological fluid and enhancing trace-level analysis reliability.
β-淀粉样蛋白（Aβ）40 与 42 之间的比例被公认为是阿尔茨海默病的生物标志物，在早期诊断和疾病进展监测中发挥着重要作用。Aβ 肽在脑脊液中的含量极低，因此，开发一种新的选择性提取方法对于从基质干扰物中分离目标生物标志物、确保准确鉴定和定量至关重要。在本研究中，在 530 µm 内径的毛细管中合成了一种有机 - 二氧化硅混合整体柱，并用于 β-淀粉样肽适配体的共价接枝。将所得的小型化寡吸附剂（mOS）应用于从人工脑脊液（CSF）样品中选择性提取 Aβ 肽 40 和 42。固定化过程实现了高于 90% 的接枝产率，导致 Aβ 适配体的密集覆盖（655 ± 15 pmol·µL 的寡吸附剂，n = 3，RSD = 2.3），且容量超过 50 pmol·µL 的 mOS。在纯介质中优化后，该 mOS 对 Aβ40 和 Aβ42 肽的提取回收率分别达到 74% 和 31%。所开发的方法包括在 mOS 上的提取和 LC-MS 分析，实现了 0.1 ng·mL 的定量下限值，精密度和准确度的 CV 和 RSD 值分别在 1.0% 至 12.9% 和 -4.7% 至 11.1% 之间。该方法成功应用于从人工脑脊液样品中选择性提取 Aβ 肽，有效地从这种复杂的生物流体中分离出两种 Aβ 目标肽，并提高了痕量水平分析的可靠性。
