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Label-free, single protein detection on a near-infrared fluorescent single-walled carbon nanotube/protein microarray fabricated by cell-free synthesis

By ?????????

Abstract

Excessive sample volumes continue to be a major limitation in the analysis of protein-protein interactions, motivating the search for label-free detection methods of greater sensitivity. Herein, we report the first chemical approach for selective protein recognition using fluorescent single-walled carbon nanotubes (SWNTs) enabling label-free microarrays capable of single protein detection. Hexahistidine-tagged capture proteins directly expressed by cell-free synthesis on SWNT/chitosan microarray are bound to a Ni2+ chelated by N??,N??-bis(carboxymethyl)-l-lysine grafted to chitosan surrounding the SWNT. The Ni2+ acts as a proximity quencher with the Ni 2+/SWNT distance altered upon docking of analyte proteins. This ability to discern single protein binding events decreases the apparent detection limit from 100 nM, for the ensemble average, to 10 pM for an observation time of 600 s. This first use of cell-free synthesis to functionalize a nanosensor extends this method to a virtually infinite number of capture proteins. To demonstrate this, the SWNT microarrays are used to analyze a network of 1156 protein-protein interactions in the staurosporine-induced apoptosis of SH-SY5Y cells, confirming literature predictions. ?? 2011 American Chemical Society

Topics: Carboxymethyl, Cell-free, chitosan, particle size, carbon nanotube, Staurosporine, Humans, Protein microarray, Biochemistry, Analytes, Proteins, Infinite numbers, fluorescence, Surface Properties, protein, structure activity relation, nanotechnology, Ensemble averages, Single-walled carbon nanotubes (SWCN), Protein detection, Microarrays, Near Infrared, Protein-protein interactions, article, Particle Size, surface property, Nanotubes, Carbon, Cell Line, Tumor, Label free, Chemical detection, Chitosan, l-Lysine, Single-walled carbon, staurosporine, Protein recognition, Sample volume, Fluorescence, Amino acids, apoptosis, drug effect, Biosynthesis, Structure-Activity Relationship, tumor cell line, Label-free detection, Detection limits, human, Protein binding, chemistry, nickel, protein microarray, protein binding, Label-free detection method, Cell death, Protein Array Analysis, Protein Binding, Nickel, Nanotechnology, Apoptosis
Publisher: American Chemical Society
Year: 2011
DOI identifier: 10.1021/nl201033d
OAI identifier: oai:repository.hanyang.ac.kr:20.500.11754/93768
Provided by: HANYANG Repository
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