Rivnak Andrew J, Rissin David M, Kan Cheuk W, Song Linan, Fishburn Matthew W, Piech Tomasz, Campbell Todd G, DuPont Derek R, Gardel Melissa, Sullivan Sean, Pink Brian A, Cabrera Carlos G, Fournier David R, Duffy David C
Quanterix Corporation, 113 Hartwell Avenue, Lexington, MA 02421, USA.
Quanterix Corporation, 113 Hartwell Avenue, Lexington, MA 02421, USA.
J Immunol Methods. 2015 Sep;424:20-7. doi: 10.1016/j.jim.2015.04.017. Epub 2015 May 7.
We report a system and assay for performing fully-automated measurement of 6 proteins simultaneously with single molecule sensitivity. The system combines handling of samples, reagents, and consumables, with a module for imaging single molecule arrays (Simoa) to enable immunoassays that have high sensitivity (~fg/mL), are multiplexed, and are fully-automated. A 6-plex cytokine Simoa assay for IL-6, TNF-α, GM-CSF, IL-10, IL-1β, and IL-1α was developed on the system. The assays had limits of detection in the range 0.01-0.03pg/mL, and the average imprecision (CV) of the Simoa signal was 4.2%. This assay was used to measure the concentrations of these cytokines in the plasma of patients with Crohn's Disease (CD), before and after treatment with anti-TNF-α antibody drugs, and in the serum of Type 1 diabetics. Concentrations of TNF-α and IL-6 in the CD samples determined using the fully-automated, multiplex Simoa assay had good correlation with the manual, single-plex assays previously reported. Drug treatment caused reductions in the mean concentration of all 6 cytokines in the plasma of CD patients. The concentrations of 4 cytokines were significantly higher in diabetics compared to healthy controls. The system could enable the widespread, multiplexed measurement of protein biomarkers with low abundance.
我们报告了一种能够以单分子灵敏度同时对6种蛋白质进行全自动测量的系统和检测方法。该系统将样品、试剂和耗材的处理与一个用于成像单分子阵列（Simoa）的模块相结合，以实现具有高灵敏度（~fg/mL）、可多重检测且全自动的免疫测定。在该系统上开发了一种用于检测白细胞介素-6（IL-6）、肿瘤坏死因子-α（TNF-α）、粒细胞-巨噬细胞集落刺激因子（GM-CSF）、白细胞介素-10（IL-10）、白细胞介素-1β（IL-1β）和白细胞介素-1α（IL-1α）的6重细胞因子Simoa检测方法。这些检测方法的检测限在0.01 - 0.03pg/mL范围内，Simoa信号的平均不精密度（CV）为4.2%。该检测方法用于测量克罗恩病（CD）患者在接受抗TNF-α抗体药物治疗前后血浆中这些细胞因子的浓度，以及1型糖尿病患者血清中这些细胞因子的浓度。使用全自动多重Simoa检测方法测定的CD样品中TNF-α和IL-6的浓度与先前报道的手工单重检测方法具有良好的相关性。药物治疗使CD患者血浆中所有6种细胞因子的平均浓度降低。与健康对照相比，糖尿病患者中4种细胞因子的浓度显著更高。该系统能够实现对低丰度蛋白质生物标志物的广泛多重测量。
