Presently, there are strong demands for libraries of small peptides containing sequences of eight residues or less for applications such as screening antibodies and specific binding proteins. Light-directed synthesis on microarray platform developed by Affymetrix offers a mean to synthesize thousands of predetermined sequence oligopeptides at defined sites. In this technique, solid-phase peptide synthesis and photolithography approach are combined to construct the peptide microarray. Light illumination through photolithographic masks is used to control the pattern of the localized photodeprotection in different areas. As a result, the simultaneous synthesis of different peptide analogs at different locations can be achieved. However, many photolithographic masks, which are very expensive to make, are usually required for the synthesis. Consequently, establishment of the method as routine is simply not possible. Therefore, the objective of this research is to develop an inexpensive method to make the peptide microarray with high efficiency and accuracy. The development of the new technique stems from two main concepts. First, an acid compound generated from triarylsulfonium hexafluoroantimonate (SSb) by light activation is used in the deprotection step of the peptide synthesis. Second, laser scanner is used to replace the photolithographic mask in order to control the locations of the illumination by laser diode to simultaneously synthesize different oligopeptides on the microwell array platform. The comparative oligopeptide synthesis using the conventional acid, trifluoroacetic acid (TFA) and the photogenerated acid reveals that SSb can replace TFA in the pentapeptide synthesis with high efficiency (93 +/- 4%). The developed technique is used to synthesize two microarray models of human protein p53 (residue 20--25) and lead binding tetrapeptide (Glu-Cys-Glu-Glu). The oligopeptide models and their analogs are parallel synthesized in the predetermined locations of the microarrays. The specific binding of the oligopeptide models is detected at the correct synthesis locations as designed and can be distinguished from their analogs. The cost of making the peptide microarray using the developed technique and the photolithographic mask technique are estimated and compared. The comparison shows that the cost to operate the deprotection of 20 amino acids using the laser scanner is less expensive than that using the photomask. The technique developed in this research can be used to make the oligopeptide microarray with high efficiency, accuracy and is cost effective. Importantly, this technique is convenient to set up in any regular research laboratories.Ph.D.Applied SciencesChemical engineeringUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/129789/2/3042100.pd
