Abstract

Protein microarrays offer a global perspective on the function of expressed gene products. However, technical issues related to the stability and dynamic range of microarrays printed with purified protein have hampered their widespread adoption. Taking an alternate approach, the Nucleic Acid Programmable Protein Array (NAPPA) is constructed by spotting protein-encoding plasmid DNA at high density, in addressable fashion, on an array surface. Proteins are subsequently generated in situ just prior to experimentation using cell-free expression systems. As such, the NAPPA platform offers a unique and viable alternative that circumvents many of the inherent limitations of spotted protein arrays, enabling diverse functional protein studies including protein-small molecule, protein-protein, antigen-antibody, and protein-nucleic acid interactions. It further offers a versatile and adaptable platform amenable to a variety of capture modalities and expression systems, and, most importantly, construction of the array is accessible to any lab with an array printer and laser slide scanner. This unit is intended to provide a reference for investigators wishing to generate arrays based on this platform, and details (1) the basic construction of cDNA-based protein microarrays from DNA isolation to printing and development, (2) quality-control efforts taken to ensure the usefulness and integrity of microarray data, and (3) a particular example of the application of self-assembling protein arrays to screen for blood-borne antibody biomarkers.

© 2011 by John Wiley & Sons, Inc.