Rapid single-molecule digital detection of protein biomarkers for continuous monitoring of systemic immune disorders.
Rapid single-molecule digital detection of protein biomarkers for continuous monitoring of systemic immune disorders.
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DOI：
10.1182/blood.2019004399
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发表时间：
2020-12
期刊：
影响因子：
20.3
通讯作者：
Yujing Song;E. Sandford;Yuzi Tian;Qingtian Yin;A. Kozminski;Shiuan-Haur Su;Tao Cai;Yuxuan Ye
中科院分区：
文献类型：
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作者：
Yujing Song;E. Sandford;Yuzi Tian;Qingtian Yin;A. Kozminski;Shiuan-Haur Su;Tao Cai;Yuxuan Ye
数字化蛋白质检测具有单分子灵敏度、高精度和稳健的测量能力，因此在推进免疫诊断方面具有巨大的潜力。然而，将数字蛋白质测定转化为急性临床护理一直具有挑战性，因为它需要快速部署。在本文中，我们提出了一种用于超快速数字蛋白质生物标志物检测的技术平台，该技术平台通过在早期预平衡状态下采用免疫复合物形成事件的单分子计数。这种方法，我们称之为“预平衡数字酶联免疫吸附测定”（PEdELISA），可以在15-300秒的前所未有的测定孵育时间内在104动态范围内定量10微升血清中的蛋白质生物标志物的多重组。PEdELISA使我们能够在表现出嵌合抗原受体T细胞（CAR-T）治疗后细胞因子释放综合征（CRS）的患者中快速监测蛋白质生物标志物，样品至应答时间约为30分钟，检测限（LOD）低于pg/mL。通过PEdELISA实现的快速、灵敏和低输入体积的生物标志物定量广泛适用于及时监测急性疾病，可能实现更个性化的治疗。
Digital protein assays have great potential to advance immunodiagnostics because of their single-molecule sensitivity, high precision, and robust measurements. However, translating digital protein assays to acute clinical care has been challenging because it requires their deployment with a rapid turnaround. Herein, we present a technology platform for ultra-fast digital protein biomarker detection by employing single-molecule counting of immune-complex formation events at an early, pre-equilibrium state. This method, which we term "pre-equilibrium digital enzyme-linked immunosorbent assay" (PEdELISA), can quantify a multiplexed panel of protein biomarkers in 10 µL of serum within an unprecedented assay incubation time of 15-300 sec over a 104 dynamic range. PEdELISA allowed us to perform rapid monitoring of protein biomarkers in patients manifesting post-chimeric antigen receptor T-cell (CAR-T) therapy cytokine release syndrome (CRS), with ~30 min sample-to-answer time and a sub-pg/mL limit of detection (LOD). The rapid, sensitive, and low input volume biomarker quantification enabled by PEdELISA is broadly applicable to timely monitoring of acute disease, potentially enabling more personalized treatment.