A Droplet Microarray for Homogeneous Enzymatic Assay

This work describes a droplet microarray for monitoring of enzymatic activity on a glass surface. These array are composed of features defined and separated by differential surface tension (surface tension array). A photolithographic method was used to create a series of circular hydrophilic spots delimited by a perfluorosilanated surface. The perfluorinated region had water contact angle on the order of 105°, whereas inside the spots this angle was < 75°. The difference in surface tension “holds” water within the hydrophilic spots thus resulting in droplet microarray. We show that each droplet can be used as a micro-reservoir for conducting of enzymatic reactions. As a proof of concept a proteolytic cleavage of a fluorogenic substrate was carried out within hundreds of confined reservoirs created on a glass slide. Using piezoelectric deposition of substrate and protease numerous reaction conditions could be tested in parallel. This format may be a valuable tool for highthroughput screening, and enzymatic profiling of novel proteins.