High Throughput Digital Droplet Assays for Ultrasensitive Multimodal (DNA, RNA, and Protein) Diagnostics

Abstract Digital assays — in which ultra-sensitive molecular measurements are made by performing millions of parallel experiments in picoliter droplets — have generated much recent enthusiasm due to their single molecule resolution of RNA, DNA, and proteins, and their robustness to reaction conditions. These assays have enormous potential for the diagnosis of difficult to diagnose diseases, such as pancreatic cancer, but are currently confined to laboratory settings due to the cumbersome instrumentation necessary to generate, control, and measure tens of millions of independent droplets. To overcome this challenge, we are developing a hybrid microelectronic / microfluidic chip to ‘unlock’ droplet-based assays for clinical use. Our microdroplet megascale detector (µMD) can generate and detect the fluorescence of millions of droplets per second (1000× faster than existing digital approaches), while achieving a 1000x greater sensitivity than conventional ELISA or ddPCR, using only a conventional cell phone camera. The key innovation of our approach is borrowed from the telecommunications industry, wherein we modulate the excitation light with a pseudorandom sequence that enables individual droplets to be resolved that would otherwise overlap due to the limited frame rate of digital cameras. Building on the success of our R21, we propose to develop a platform technology to ultrasensitively quantify proteins, ctDNA, and single EVs in an integrated device, directly in patient blood, to address critical issues in multimodal diagnostics. In collaboration with the Abramson Cancer Center and building on prior work together on multi-modal diagnostics, we focus our attention on Pancreatic ductal adenocarcinoma-the third leading cause of cancer- related death in the United States with an overall 5-year survival of only 9%.