Protein Microarray Copying: Easy on‐Demand Protein Microarray Generation Compatible with Fluorescence and Label‐Free Real‐Time Analysis

다국어 초록 (Multilingual Abstract)

Protein microarrays are essential to understand complex protein interaction networks. Their production, however, is a challenge and renders this technology unattractive for many laboratories. Recent developments in cell‐free protein microarray generation offer new opportunities, but are still expensive and cumbersome in practice. Herein, we describe a cost‐effective and user‐friendly method for the cell‐free production of protein microarrays. From a polydimethylsiloxane (PDMS) flow cell containing an expressible DNA microarray, proteins of interest are synthesised by cell‐free expression and then immobilised on a capture surface. The resulting protein microarray can be regarded as a “copy” of the DNA microarray. 2 His6‐ and Halo‐tagged fluorescent reference proteins were used to demonstrate the functionality of nickel nitrilotriacetic acid (Ni‐NTA) and Halo‐bind surfaces in this copy system. The described process can be repeated several times on the same DNA microarray. The identity and functionality of the proteins were proven during the copy process by their fluorescence and on the surface through a fluorescent immune assay. Also, single‐colour reflectometry (SCORE) was applied to show that, on such copied arrays, real‐time binding kinetic measurements were possible.
One‐hour copying: A protein microarray can be generated on demand within an hour, with cost‐effective and low‐tech equipment. This is achieved by “copying” DNA on a microfluidic flow cell into proteins in a process called protein microarray copying. Different protein‐capture surfaces have been validated for this technique. Label‐free detection of protein–protein interactions has also been applied to these copied protein microarrays.