Procalcitonin detection in human plasma specimens using a fast version of proximity extension assay.

Proximity Extension Assay (PEA), an homogeneous, dual-recognition immunoassay, has proven to be sensitive, specific and convenient for detection or quantitation of one or multiple analytes in human plasma. In this paper, the PEA principle was applied to the detection of procalcitonin (PCT), a widely used biomarker for the identification of bacterial infection. A simple, short PEA protocol, with an assay time suitable for point-of-care diagnostics, is presented here as a proof of concept. Pairs of oligonucleotides and monoclonal antibodies were selected to generate tools specifically adapted to the development of an efficient PEA for PCT detection. The assay time was reduced by more than 13-fold compared to published versions of PEA,  without significantly affecting assay performance. It was also demonstrated that T4 DNA polymerase could advantageously be replaced by other polymerases having strong 3’>5’ exonuclease activity. The sensitivity of this improved assay was determined to be about 0.1 ng/mL of PCT in plasma specimen.

邻近延伸测定法（Proximity Extension Assay, PEA）作为一种均相双识别免疫测定技术，已被证实可灵敏、特异且便捷地检测或定量人血浆中的单种或多种分析物。本文将PEA原理应用于降钙素原（procalcitonin, PCT）的检测——PCT是目前广泛用于细菌感染鉴别诊断的经典生物标志物。本研究报道了一种简便快捷的PEA检测流程，其测定时长适配即时诊断场景，并完成了概念验证：我们筛选得到适配降钙素原高效PEA检测的寡核苷酸与单克隆抗体工具组合。相较于已发表的PEA检测方案，本方法将测定时长缩短了13倍以上，且未显著影响测定性能。研究同时证实，可使用具备强3’→5’核酸外切酶活性的其他聚合酶，替代传统的T4 DNA聚合酶以获得更优效果。经测定，该优化后的检测方法对血浆样本中降钙素原的检测灵敏度约为0.1 ng/mL。