Abstract Traditionally, protein analytes or biomarkers have been measured individually in ELISAs. However, it is becoming increasingly clear that multiple markers are associated with multivariate diseases like cancer. Despite demand, the adoption of multi-analyte biomarker tests in clinical research has been severely limited for many reasons including technical concerns regarding assay reproducibility, time and the labor-intensive nature of assay panel development, and reported non-correlation with conventional ELISA data. To address this issue, we have developed a novel fully automated desktop immunoassay platform that enables simultaneous interrogation of four analytes in eight individual samples on a single disposable microfluidic cartridge in under an hour. We have devised a unique approach to immobilize antibodies that results in assays with high sensitivities (femtomolar) and broad dynamic ranges (>4 logs). Twenty microliters of patient sample are split into parallel channels containing individual immunoassays. By segregating each assay to a discrete location, the multi-analyte assay mimics single-analyte ELISAs, thus enabling each assay to operate under its optimal conditions. This methodology combines the specificity of a traditional ELISA immunoassay, where the sample is assayed by a singular pair of capture and detect antibodies eliminating cross-reactivity and other negative interactions from other antibody pairs while offering the benefits of a multiplexed antigen analysis. We describe the system's design, speed, sensitivity and reproducibility using data generated with a custom-built 4-analyte pro-inflammatory panel cartridge (IL-1β, IL-6, IL-10 and TNFα) on clinical samples. We demonstrate that our assay system is ideal for quantitative analysis of soluble protein markers in clinical research and in clinical trials. Citation Format: Rajiv Pande, John H. Leamon, Marty Putnam, M Pitta, D Svancara, J Tu. A microfluidic, multi-analyte, high sensitivity, immunoassay platform for rapid biomarker detection. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3220. doi:10.1158/1538-7445.AM2013-3220
Read full abstract