title: High-throughput screening in two dimensions: binding intensity and off-rate on a peptide microarray.
abstract: We report a high-throughput two-dimensional microarray-based screen, incorporating both target binding intensity and off-rate, which can be used to analyze thousands of compounds in a single binding assay. Relative binding intensities and time-resolved dissociation are measured for labeled tumor necrosis factor alpha (TNF-alpha) bound to a peptide microarray. The time-resolved dissociation is fitted to a one-component exponential decay model, from which relative dissociation rates are determined for all peptides with binding intensities above background. We show that most peptides with the slowest off-rates on the microarray also have the slowest off-rates when measured by surface plasmon resonance (SPR).
authors.affiliation: The Biodesign Institute at Arizona State University, 1001 S. McAllister Avenue, Tempe, AZ 85287-5201, USA.
authors.forename: Matthew P
authors.initials: MP
authors.lastname: Greving
authors.forename: Paul E
authors.initials: PE
authors.lastname: Belcher
authors.forename: Conor D
authors.initials: CD
authors.lastname: Cox
authors.forename: Douglas
authors.initials: D
authors.lastname: Daniel
authors.forename: Chris W
authors.initials: CW
authors.lastname: Diehnelt
authors.forename: Neal W
authors.initials: NW
authors.lastname: Woodbury
mesh_terms: D000595:Amino Acid Sequence; D057166:High-Throughput Screening Assays / Q000379:methods; D008969:Molecular Sequence Data; D019151:Peptide Library; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D040081:Protein Array Analysis / Q000379:methods*; D011485:Protein Binding; D020349:Surface Plasmon Resonance / Q000379:methods*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*
chemical_list: D019151:Peptide Library; D010455:Peptides; D014409:Tumor Necrosis Factor-alpha