title: Aptamer-based organic-silica hybrid affinity monolith prepared via "thiol-ene" click reaction for extraction of thrombin.
abstract: A novel strategy for preparing aptamer-based organic-silica hybrid monolithic column was developed via "thiol-ene" click chemistry. Due to the large specific surface area of the hybrid matrix and the simplicity, rapidness and high efficiency of "thiol-ene" click reaction, the average coverage density of aptamer on the organic-silica hybrid monolith reached 420 pmol μL(-1). Human α-thrombin can be captured on the prepared affinity monolithic column with high specificity and eluted by NaClO4 solution. N-p-tosyl-Gly-Pro-Arg p-nitroanilide acetate was used as the sensitive chromogenic substrate of thrombin. The thrombin enriched by this affinity column was detected with a detection of limit of 0.01 μM by spectrophotometry. Furthermore, the extraction recovery of thrombin at 0.15 μM in human serum was 91.8% with a relative standard deviation of 4.0%. These results indicated that "thiol-ene" click chemistry provided a promising technique to immobilize aptamer on organic-inorganic hybrid monolith and the easily-assembled affinity monolithic material could be used to realize highly selective recognition of trace proteins.
authors: State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry & Chemical Engineering and Center of Materials Analysis, Nanjing University, 22 Hankou Road, Nanjing 210093, China.
authors: Zheng
authors: Z
authors: Wang
authors: State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry & Chemical Engineering and Center of Materials Analysis, Nanjing University, 22 Hankou Road, Nanjing 210093, China.
authors: Jin-Cheng
authors: JC
authors: Zhao
authors: State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry & Chemical Engineering and Center of Materials Analysis, Nanjing University, 22 Hankou Road, Nanjing 210093, China. Electronic address: hzlian@nju.edu.cn.
authors: Hong-Zhen
authors: HZ
authors: Lian
authors: State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry & Chemical Engineering and Center of Materials Analysis, Nanjing University, 22 Hankou Road, Nanjing 210093, China.
authors: Hong-Yuan
authors: HY
authors: Chen
mesh_terms: D052158:Aptamers, Peptide / Q000737:chemistry*; D002853:Chromatography, Liquid; D057930:Click Chemistry / Q000379:methods*; D006801:Humans; D009842:Oligopeptides / Q000737:chemistry*; D044967:Serum / Q000737:chemistry*; D012822:Silicon Dioxide / Q000737:chemistry*; D013438:Sulfhydryl Compounds / Q000737:chemistry*; D013917:Thrombin / Q000032:analysis* / Q000302:isolation & purification*
chemical_list: D052158:Aptamers, Peptide; D009842:Oligopeptides; D013438:Sulfhydryl Compounds; C035024:glycyl-prolyl-arginine; D012822:Silicon Dioxide; D013917:Thrombin
keywords: Aptamer; Enrichment; Enzyme reaction; Hybrid monolithic column; “Thiol-ene” click chemistry