基于雜交鏈?zhǔn)椒磻?yīng)檢測赭曲霉毒素A的高效電化學(xué)適體傳感器研究
發(fā)布時間：2018-10-15 14:51
【摘要】：赭曲霉毒素(Ochratoxins,OTs)是一種由赭曲霉、碳黑曲霉和純綠青霉產(chǎn)生的食源性毒素,主要污染糧谷類農(nóng)產(chǎn)品,與人類和動物健康關(guān)系密切。其中以赭曲霉毒素A(Ochratoxin A,OTA)的毒性最大、散布最廣、污染程度最高。本研究利用電化學(xué)適體傳感器基于雜交鏈?zhǔn)椒磻?yīng)研發(fā)設(shè)計了一種擁有超低檢測限的OTA檢測方法。首先,將一段捕獲探針固定在金電極上,然后加入與之互補(bǔ)的OTA適配體序列,使其與捕獲探針形成DNA雜交鏈。在環(huán)境中存在OTA的情況下,雜交鏈上的OTA適配體會與游離的OTA結(jié)合,脫離電極。然后,加入檢測探針和兩段生物素標(biāo)記的互補(bǔ)DNA發(fā)夾結(jié)構(gòu),與電極上剩余的捕獲探針結(jié)合,并引發(fā)雜交鏈?zhǔn)椒磻?yīng)。最后,在電極上滴加堿性磷酸酶(ST-AP),它與DNA鏈上的生物素結(jié)合,通過催化反應(yīng)底物中的-萘酚磷酸鹽(-NP)產(chǎn)生電化學(xué)信號,DPV曲線隨OTA濃度的增高而增高。在優(yōu)化條件下,該檢測平臺對OTA的檢測線性范圍是0.005-100ng/m L,檢測限可達(dá)2 pg/m L。此外,該方法在檢測谷物樣品的OTA應(yīng)用中,也獲得了較為滿意的回收率,檢測結(jié)果與高效液相色譜法(HPLC)相比具有高度的一致性。
[Abstract]:Ochratoxin (Ochratoxins,OTs) is a foodborne toxin produced by Aspergillus ochratus, Aspergillus Niger and Pure Penicillium. It mainly pollutes cereals and is closely related to human and animal health. OTA of ochratoxin A (Ochratoxin was the most toxic, most widespread and the most polluted. In this study, an ultra-low detection limit (OTA) detection method based on hybrid chain reaction was developed using electrochemical aptamer sensor. Firstly, a capture probe was immobilized on the gold electrode, and then the complementary OTA aptamer sequence was added to form the DNA hybridization chain with the capture probe. In the presence of OTA in the environment, the adaptation of OTA on the hybrid chain was associated with the free OTA, and the electrode was detached. Then, the complementary DNA hairpin structure was added with the detection probe and biotinylated probe to combine with the remaining trapping probe on the electrode, and the hybridization chain reaction was initiated. Finally, alkaline phosphatase (ST-AP) was added to the electrode, which binds to the biotin in the DNA chain and produces electrochemical signals by catalyzing-naphthol phosphate (- NP) in the substrate. The DPV curve increases with the increase of OTA concentration. Under the optimized conditions, the linear range of the detection platform for OTA is 0.005-100ng/m L, and the detection limit can reach 2 pg/m L. In addition, the method has been applied to the determination of grain samples with OTA, and the results are in good agreement with that of (HPLC) by high performance liquid chromatography (HPLC).
【學(xué)位授予單位】：重慶醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：TP212.3;R440
本文編號：2272892
[Abstract]:Ochratoxin (Ochratoxins,OTs) is a foodborne toxin produced by Aspergillus ochratus, Aspergillus Niger and Pure Penicillium. It mainly pollutes cereals and is closely related to human and animal health. OTA of ochratoxin A (Ochratoxin was the most toxic, most widespread and the most polluted. In this study, an ultra-low detection limit (OTA) detection method based on hybrid chain reaction was developed using electrochemical aptamer sensor. Firstly, a capture probe was immobilized on the gold electrode, and then the complementary OTA aptamer sequence was added to form the DNA hybridization chain with the capture probe. In the presence of OTA in the environment, the adaptation of OTA on the hybrid chain was associated with the free OTA, and the electrode was detached. Then, the complementary DNA hairpin structure was added with the detection probe and biotinylated probe to combine with the remaining trapping probe on the electrode, and the hybridization chain reaction was initiated. Finally, alkaline phosphatase (ST-AP) was added to the electrode, which binds to the biotin in the DNA chain and produces electrochemical signals by catalyzing-naphthol phosphate (- NP) in the substrate. The DPV curve increases with the increase of OTA concentration. Under the optimized conditions, the linear range of the detection platform for OTA is 0.005-100ng/m L, and the detection limit can reach 2 pg/m L. In addition, the method has been applied to the determination of grain samples with OTA, and the results are in good agreement with that of (HPLC) by high performance liquid chromatography (HPLC).
【學(xué)位授予單位】：重慶醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：TP212.3;R440
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