High-Throughput Mutiplexed Protease Activity Measurement Using a Droplet Based Microfluidic Platform with Picoinjector

Abstract

In this study, we integrated several components, including a droplet generator, a pico-injector, and an analytical inference technique, Proteolytic Activity Matrix Analysis (PrAMA), to create a platform for assessing multiple specific protease activity assays with minimal liquid handling and sample-requirement for personal medicine analysis. The microfluidic platform enables the direct measurements of protease enzyme activity, which is more physiologically informative than the standard measurements to determine the enzyme concentration alone. By tracking hundreds of picoliter droplets containing biological samples mixed with unique FRET-based protease substrates and inhibitors, the assay simultaneously infers multiple specific protease activities with minimal (<20µl) physiological sample requirement. We applied this method to in vitro study of an immortalized cell line established from a peritoneal endometriotic biopsy to ascertain the proteolytic activity response of TNF-α treatments.