Menero-Valdés Paula, Lores-Padín Ana, Fernández Beatriz, González-Iglesias Héctor, Pereiro Rosario
Department of Physical and Analytical Chemistry, University of Oviedo, Julian Claveria 8, 33006, Oviedo, Spain.
Department of Physical and Analytical Chemistry, University of Oviedo, Julian Claveria 8, 33006, Oviedo, Spain; Instituto Universitario Fernández-Vega, Fundación de Investigación Oftalmológica, Universidad de Oviedo, Oviedo, Spain.
Talanta. 2022 Jul 1;244:123424. doi: 10.1016/j.talanta.2022.123424. Epub 2022 Mar 29.
Iridium nanoclusters (IrNCs) stabilized with citrate were synthesized and then the ligand was exchanged by lipoic acid (LA). The IrNCs@LA were bioconjugated through carbodiimide chemistry with specific antibodies to prepare IrNCs-labelled immunoprobes. The IrNCs-immunoprobes were employed in competitive immunoassays for immunoglobulin E (IgE) and apolipoprotein E (APOE) determination by detecting the iridium through inductively coupled plasma - mass spectrometry (ICP-MS). The IrNCs@LA have a 1.89 nm diameter at average and each NC contains 250 Ir atoms. Labelling of specific antibodies with IrNCs was optimized in terms of recognition capabilities and signal amplification by ICP-MS. Amplification and detection limits can be tuned by selecting the IrNCs:Ab molar ratio. An immunoprobe prepared by mixing a 10:1 IrNC:Ab molar ratio was selected for the determination of IgE and APOE in aqueous humor, achieving a signal amplification of 1760 iridium atoms per molecule of the sought protein and limits of detection in the tens of pg mL of protein. The IrNCs-immunoprobes were evaluated for IgE determination in serum samples as well as for IgE and APOE in aqueous humor (from controls subjects and patients affected by primary open angle glaucoma) by ICP-MS, being required just sample dilution as pre-treatment. Results were corroborated with commercial ELISA kits.
合成了用柠檬酸盐稳定的铱纳米团簇（IrNCs），然后用硫辛酸（LA）交换配体。通过碳二亚胺化学将IrNCs@LA与特异性抗体进行生物偶联，以制备IrNCs标记的免疫探针。通过电感耦合等离子体质谱（ICP-MS）检测铱，将IrNCs免疫探针用于免疫球蛋白E（IgE）和载脂蛋白E（APOE）的竞争性免疫测定。IrNCs@LA的平均直径为1.89纳米，每个纳米团簇包含250个铱原子。就识别能力和ICP-MS信号放大而言，对用IrNCs标记特异性抗体进行了优化。通过选择IrNCs:Ab摩尔比可以调节放大倍数和检测限。选择通过混合10:1的IrNC:Ab摩尔比制备的免疫探针来测定房水中的IgE和APOE，每分子目标蛋白实现了1760个铱原子的信号放大，蛋白质检测限在数十pg/mL。通过ICP-MS评估IrNCs免疫探针用于血清样品中IgE的测定以及房水中（来自对照受试者和原发性开角型青光眼患者）IgE和APOE的测定，仅需将样品稀释作为预处理。结果与商用ELISA试剂盒得到了证实。
