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微流控数字免疫分析用于即时检测全血中的 NT-proBNP。

Microfluidic Digital Immunoassay for Point-of-Care Detection of NT-proBNP from Whole Blood.

机构信息

School of Biological and Health Systems Engineering, Arizona State University, Tempe, Arizona 85287, United States.

Center for Biosensors and Bioelectronics, The Biodesign Institute, Arizona State University, Tempe, Arizona 85287, United States.

出版信息

Anal Chem. 2024 Jul 2;96(26):10569-10576. doi: 10.1021/acs.analchem.4c01046. Epub 2024 Jun 15.

Abstract

The high prevalence and economic burden of heart failure remain a challenge to global health. This lifelong disease leads to a buildup of permanent heart damage, making early detection and frequent monitoring crucial for effective treatment. N-terminal proBNP (NT-proBNP) is an important biomarker for monitoring the disease state, but current commercial and research NT-proBNP assays require phlebotomy and bulky equipment or do not satisfy clinical requirements such as sensitivity and detection thresholds. Here, we report a point-of-care (POC) compatible microfluidic digital immunoassay that can quantify the NT-proBNP concentration in a small volume of whole blood. Our automated microfluidic device takes whole blood samples mixed with biotinylated detection antibodies and passes through a plasma filter to react with a capture antibody-functionalized sensor surface. Streptavidin-coated gold nanoparticles (GNPs) are then released to mark the surface-bound single NT-proBNP immunocomplexes and recorded with bright-field microscopy. NT-proBNP concentrations in the sample are quantified via a hybrid digital/analog calibration curve. Digital counts of bound GNPs are used as readout signal at low concentrations for high sensitivity detection, and GNP pixel occupancies are used at high concentrations for extended dynamic range. With this approach, we detected NT-proBNP in the range of 8.24-10 000 pg/mL from 7 μL of whole blood in 10 min, with a limit of detection of 0.94 pg/mL. Finally, the method was validated with 15 clinical serum samples, showing excellent linear correlation ( = 0.998) with Roche's Elecsys proBNP II assay. This evidence indicates that this method holds promise for decentralized monitoring of heart failure.

摘要

心力衰竭的高患病率和经济负担仍然是全球健康的一大挑战。这种终身疾病会导致永久性心脏损伤的积累,因此早期检测和频繁监测对于有效治疗至关重要。N 端脑利钠肽前体(NT-proBNP)是监测疾病状态的一个重要生物标志物,但目前的商业和研究用 NT-proBNP 检测需要采血和使用笨重的设备,或者不能满足灵敏度和检测阈值等临床要求。在这里,我们报告了一种即时检测(POC)兼容的微流控数字免疫分析方法,可用于定量检测小体积全血中的 NT-proBNP 浓度。我们的自动化微流控设备采用全血样本,与生物素化检测抗体混合,然后通过血浆过滤器与固定在传感器表面的捕获抗体发生反应。然后,链霉亲和素包被的金纳米颗粒(GNPs)被释放出来,标记表面结合的单个 NT-proBNP 免疫复合物,并通过明场显微镜记录。通过混合数字/模拟校准曲线来定量样品中的 NT-proBNP 浓度。在低浓度下,结合的 GNPs 的数字计数用作读出信号,以实现高灵敏度检测,而在高浓度下,GNP 像素占有率用于扩展动态范围。通过这种方法,我们在 10 分钟内从 7 μL 全血中检测到 8.24-10000 pg/mL 的 NT-proBNP,检测限为 0.94 pg/mL。最后,该方法用 15 份临床血清样本进行了验证,与罗氏的 Elecsys proBNP II 检测法具有极好的线性相关性( = 0.998)。这些证据表明,该方法有望用于心力衰竭的分散式监测。

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