Figure 4.2.

Fabrication of high-content functional protein microarrays.

Four major steps are involved to construct a functional protein microarray of high content. First, a high-quality ORF expression library is constructed to allow inducible overexpression of GST-His6 fused recombinant proteins in yeast. Second, a high-throughput protein purification protocol is applied to individually purify thousands of proteins from yeast. The purified proteins are stored in 384-well format. Third, silver stain and immunoblot analysis are employed to evaluate the quality and quantity of the purified proteins. Finally, when purified proteins pass the quality control, they are spotted in duplicate to glass slides using a robot microarrayer. The quality of printing is then tested by anti-GST probing. An image of a human protein microarray probed with anti-GST is shown in the lower panel. For color version of this figure, the reader is referred to the online version of this book.