title: High density diffusion-free nanowell arrays.
abstract: Proteomics aspires to elucidate the functions of all proteins. Protein microarrays provide an important step by enabling high-throughput studies of displayed proteins. However, many functional assays of proteins include untethered intermediates or products, which could frustrate the use of planar arrays at very high densities because of diffusion to neighboring features. The nucleic acid programmable protein array (NAPPA) is a robust in situ synthesis method for producing functional proteins just-in-time, which includes steps with diffusible intermediates. We determined that diffusion of expressed proteins led to cross-binding at neighboring spots at very high densities with reduced interspot spacing. To address this limitation, we have developed an innovative platform using photolithographically etched discrete silicon nanowells and used NAPPA as a test case. This arrested protein diffusion and cross-binding. We present confined high density protein expression and display, as well as functional protein-protein interactions, in 8000 nanowell arrays. This is the highest density of individual proteins in nanovessels demonstrated on a single slide. We further present proof of principle results on ultrahigh density protein arrays capable of up to 24000 nanowells on a single slide.
authors: Biodesign Institute, Center for Personal Diagnostics, Arizona State University, Tempe, Arizona 85287, United States.
authors: Bharath R
authors: BR
authors: Takulapalli
authors: Ji
authors: J
authors: Qiu
authors: D Mitchell
authors: DM
authors: Magee
authors: Peter
authors: P
authors: Kahn
authors: Al
authors: A
authors: Brunner
authors: Kristi
authors: K
authors: Barker
authors: Steven
authors: S
authors: Means
authors: Shane
authors: S
authors: Miersch
authors: Xiaofang
authors: X
authors: Bian
authors: Alex
authors: A
authors: Mendoza
authors: Fernanda
authors: F
authors: Festa
authors: Karan
authors: K
authors: Syal
authors: Jin G
authors: JG
authors: Park
authors: Joshua
authors: J
authors: LaBaer
authors: Peter
authors: P
authors: Wiktor
mesh_terms: D004058:Diffusion; D006801:Humans; D056656:Lab-On-A-Chip Devices; D040081:Protein Array Analysis / Q000295:instrumentation*; D014176:Protein Biosynthesis; D025941:Protein Interaction Mapping; D020543:Proteome / Q000096:biosynthesis / Q000235:genetics; D040901:Proteomics; D012825:Silicon / Q000737:chemistry
chemical_list: D020543:Proteome; D012825:Silicon