Liquid Crystal Multiplexed Protease Assays Reporting Enzymatic Activities as Optical Bar Charts

We describe a highly sensitive, substrate-specific, label-free, and multiplexed protease assay which reports proteases activities as an optical bar chart, allowing test results to be easily assessed by laymen with the naked eye. First, an oligopeptide microarray having six rows of immobilized oligopeptides, with well-controlled orientations and concentration gradients, is immersed in a buffer solution containing proteases. Then, a thin layer liquid crystal is supported on the microarray to transduce the oligopeptide cleavage event into an optical bar chart of different colors and lengths. This type of optical bar chart provides very rich information such as protease concentration, incubation time, surface densities of oligopeptides, etc. Both trypsin and chymotrypsin can be detected by using this assay within 3 h. The capability of the multiplexed protease assay opens up possibilities for detecting toxins such as botulinum neurotoxins which are known to cleave proteins and affect the docking and fusing synaptic vesicles.