A highly sensitive fluorescence resonance energy transfer aptasensor for staphylococcal enterotoxin B detection based on exonuclease-catalyzed target recycling strategy

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• 作者：Shijia Wu ; Nuo Duan ; Xiaoyuan Ma ; Yu Xia ; Hongxin Wang ; Zhouping Wang ; ^{wangzp@jiangnan.edu.cn}
• 关键词：Fluorescence resonance energy transfer ; Staphylococcal enterotoxin B ; Mn2+ doped NaYF4 ; Yb/Er upconversion nanoparticles ; Exonuclease-catalyzed
• 刊名：Analytica Chimica Acta
• 出版年：2013
• 出版时间：11 June, 2013
• 年：2013
• 卷：782
• 期：Complete
• 页码：59-66
• 全文大小：1295 K

文摘

An ultrasensitive fluorescence resonance energy transfer (FRET) bioassay was developed to detect staphylococcal enterotoxin B (SEB), a low molecular exotoxin, using an aptamer-affinity method coupled with upconversion nanoparticles (UCNPs)-sensing, and the fluorescence intensity was prominently enhanced using an exonuclease-catalyzed target recycling strategy. To construct this aptasensor, both fluorescence donor probes (complementary DNA₁-UCNPs) and fluorescence quencher probes (complementary DNA₂-Black Hole Quencher₃ (BHQ₃)) were hybridized to an SEB aptamer, and double-strand oligonucleotides were fabricated, which quenched the fluorescence of the UCNPs via FRET. The formation of an aptamer-SEB complex in the presence of the SEB analyte resulted in not only the dissociation of aptamer from the double-strand DNA but also both the disruption of the FRET system and the restoration of the UCNPs fluorescence. In addition, the SEB was liberated from the aptamer-SEB complex using exonuclease I, an exonuclease specific to single-stranded DNA, for analyte recycling by selectively digesting a particular DNA (SEB aptamer). Based on this exonuclease-catalyzed target recycling strategy, an amplified fluorescence intensity could be produced using different SEB concentrations. Using optimized experimental conditions produced an ultrasensitive aptasensor for the detection of SEB, with a wide linear range of 0.001-1 ng mL^?1 and a lower detection limit (LOD) of 0.3 pg mL^?1 SEB (at 3¦Ò). The fabricated aptasensor was used to measure SEB in a real milk samples and validated using the ELISA method. Furthermore, a novel aptasensor FRET assay was established for the first time using 30 mol % Mn²⁺ ions doped NaYF₄:Yb/Er (20/2 mol % ) UCNPs as the donor probes, which suggests that UCNPs are superior fluorescence labeling materials for food safety analysis.