On-Chip Protein Synthesis for Making Microarrays

Protein microarrays are a miniaturized format for displaying in close spatial density hundreds or thousands of purified proteins that provide a powerful platform for the high-throughput assay of protein function. The traditional method of producing them requires the high-throughput production and printing of proteins, a laborious method that raises concerns about the stability of the proteins and the shelf life of the arrays. A novel method of producing protein microarrays, called nucleic acid programmable protein array (NAPPA), overcomes these limitations by synthesizing proteins in situ . NAPPA entails spotting plasmid DNA encoding the relevant proteins, which are then simultaneously transcribed and translated by a cell-free system. The expressed proteins are captured and oriented at the site of expression by a capture reagent that targets a fusion protein on either the N- or C-terminus of the protein. Using a mammalian extract, NAPPA expresses and captures 1000-fold more protein per feature than conventional protein-printing arrays. Moreover, this approach minimizes concerns about protein stability and integrity, because proteins are produced just in time for assaying. NAPPA has already proven to be a robust tool for protein functional assays.