title: A simple and sensitive label-free fluorescent approach for protein detection based on a Perylene probe and aptamer.
abstract: Highly sensitive detection of proteins is of great importance for effective clinical diagnosis and biomedical research. However, so far most detection methods rely on antibody-based immunoassays and are usually laborious and time-consuming with poor sensitivity. Here, we developed a simple and ultra-sensitive method to detect a biomarker protein-thrombin by taking advantage of the fluorescent probe Perylene tetracarboxylic acid diimide (PTCDI) derivatives and thrombin aptamer. The water-soluble dye PTCDI shows strong fluorescence in buffer solution for the existence of free dye monomer, but becomes weak after aggregation through self-assembly on nucleic acid aptamer. In the presence of thrombin, it specifically binds to thrombin aptamer which causes the conformational transition between aptamer and PTCDI and results in a significant fluorescence recovery. The results showed that as low as 40 pM of thrombin could be detected by this method. The high sensitivity of the developed sensing system mainly attributes to the ultra-sensitivity of the fluorescence intensity changes of PTCDI. With the specificity of aptamer, the assay exhibited high selectivity for thrombin against three other proteins (bovine serum albumin, lysozyme, mouse IgG) and 1% diluted fetal bovine serum. The detection method might be extended to sensitive detection of a variety of proteins for its advantages of isothermal conditions required, simple and rapid without multiple separation and washing steps.
authors.affiliation: Institute of Quality Standards and Testing Technology for Agro-products, Key Laboratory of Agro-product Quality and Safety, Chinese Academy of Agricultural Sciences, Beijing 100081, China; Key Laboratory of Agri-food Quality and Safety, Ministry of Agriculture, Beijing 100081, China; College of Food Science, Sichuan Agricultural University, Ya'an 625014, China.
authors.forename: Zhenzhen
authors.initials: Z
authors.lastname: Lv
authors.affiliation: Institute of Quality Standards and Testing Technology for Agro-products, Key Laboratory of Agro-product Quality and Safety, Chinese Academy of Agricultural Sciences, Beijing 100081, China; Key Laboratory of Agri-food Quality and Safety, Ministry of Agriculture, Beijing 100081, China.
authors.forename: Jinchuan
authors.initials: J
authors.lastname: Liu
authors.affiliation: Institute of Quality Standards and Testing Technology for Agro-products, Key Laboratory of Agro-product Quality and Safety, Chinese Academy of Agricultural Sciences, Beijing 100081, China; Key Laboratory of Agri-food Quality and Safety, Ministry of Agriculture, Beijing 100081, China.
authors.forename: Wenhui
authors.initials: W
authors.lastname: Bai
authors.affiliation: Institute of Quality Standards and Testing Technology for Agro-products, Key Laboratory of Agro-product Quality and Safety, Chinese Academy of Agricultural Sciences, Beijing 100081, China; Key Laboratory of Agri-food Quality and Safety, Ministry of Agriculture, Beijing 100081, China.
authors.forename: Shuming
authors.initials: S
authors.lastname: Yang
authors.affiliation: Institute of Quality Standards and Testing Technology for Agro-products, Key Laboratory of Agro-product Quality and Safety, Chinese Academy of Agricultural Sciences, Beijing 100081, China; Key Laboratory of Agri-food Quality and Safety, Ministry of Agriculture, Beijing 100081, China. Electronic address: ailiang.chen@gmail.com.
authors.forename: Ailiang
authors.initials: A
authors.lastname: Chen
mesh_terms: D052157:Aptamers, Nucleotide / Q000032:analysis / Q000737:chemistry* / Q000235:genetics; D005456:Fluorescent Dyes / Q000737:chemistry; D010569:Perylene / Q000737:chemistry*; D040081:Protein Array Analysis / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013050:Spectrometry, Fluorescence / Q000379:methods*; D013194:Staining and Labeling; D013917:Thrombin / Q000032:analysis* / Q000737:chemistry / Q000235:genetics
chemical_list: D052157:Aptamers, Nucleotide; D005456:Fluorescent Dyes; D010569:Perylene; D013917:Thrombin
keywords: Aptamer; Fluorescence; Label-free; Perylene tetracarboxylic acid diimide; Thrombin