title: A sensitive assay for trypsin using poly(thymine)-templated copper nanoparticles as fluorescent probes.
abstract: A new, simple and sensitive fluorescence strategy was developed for the trypsin assay based on copper nanoparticles (CuNPs) and its different fluorescence response toward trypsin-catalyzed hydrolysis of cytochrome c (Cyt c). Polythymine (poly T)-templated CuNPs served as effective fluorescent probes. Cyt c is well-known to act as a quencher. However, herein, a low concentration of Cyt c was designed specially to act as the substrate of trypsin to avoid the quenching effects by electron transfer from Cyt c to CuNPs. In the presence of trypsin, Cyt c hydrolyzes to small peptides, releasing free cysteine residues. Nonfluorescent coordination complexes were formed upon exposure to free cysteine residues by a metal-ligand bond between Cu atoms and sulfur atoms, leading to a decreased fluorescence response to CuNPs. This novel method for the quantitative determination of trypsin has a linear detection range from 0.25 μg mL(-1) to 1000 μg mL(-1) and a relatively low detection limit of 42 ng mL(-1). To the best of our knowledge, this is the first application of the trypsin-catalyzed hydrolysis reaction of Cyt c to produce quenching effect in bioanalysis, which provided a novel approach for the biochemical sensing strategy.
authors: College of Material and Chemical Engineering, Hunan Institute of Technology, Hengyang 421002, P. R. China. oulijuanann@aliyun.com.
authors: Li-Juan
authors: LJ
authors: Ou
authors: Xiao-Yan
authors: XY
authors: Li
authors: Li-Juan
authors: LJ
authors: Li
authors: Hong-Wei
authors: HW
authors: Liu
authors: Ai-Ming
authors: AM
authors: Sun
authors: Kai-Jian
authors: KJ
authors: Liu
mesh_terms: D000818:Animals; D015374:Biosensing Techniques / Q000379:methods; D002417:Cattle; D003300:Copper / Q000737:chemistry*; D045304:Cytochromes c / Q000737:chemistry / Q000378:metabolism; D005456:Fluorescent Dyes / Q000737:chemistry*; D006801:Humans; D006868:Hydrolysis; D057230:Limit of Detection; D053758:Nanoparticles / Q000737:chemistry*; D011071:Poly T / Q000737:chemistry*; D013050:Spectrometry, Fluorescence / Q000379:methods; D014357:Trypsin / Q000032:analysis* / Q000378:metabolism
chemical_list: D005456:Fluorescent Dyes; D011071:Poly T; D003300:Copper; D045304:Cytochromes c; D014357:Trypsin