【摘要】：大腸桿菌0157:H7(E. coli 0157:H7)作為危害最大的食源性致病菌之一,在我國存在流行暴發(fā)的可能性和感染的嚴重性。由于傳統檢測方法存在操作繁瑣、費力、耗時等缺點,因此急需研制一種快速準確的檢測方法。本課題調查了目前國際上諸多新型檢測技術,包括生物傳感器。根據Spreeta SPR傳感器的優(yōu)勢,最終將Spreeta SPR傳感器與細胞裂解法作為主要研究手段與內容,實現對大腸桿菌O157：H7的檢測,并對其進行了較為深入的探討和研究。 本課題的主要研究內容和研究結果如下： (1)用不同濃度葡萄糖溶液評估Spreeta SPR傳感器性能,葡萄糖濃度與折射率間的相關系數接近1,用去離子水沖洗后回復基線,得出該傳感器性能良好。 (2)通過親和素及生物素標記抗體的響應結果的重復性實驗可知該傳感器重復性高,為Spreeta SPR免疫傳感器用于檢測大腸桿菌O157：H7提供了理論基礎,其檢測限為3×106 cfu mL-1,基本滿足檢測需求。 (3)比較BIACORE SPR免疫傳感器,用直接法檢測大腸桿菌O157：H7的檢測限為3×105 cfu mL-1,檢測靈敏度與酶聯免疫吸附法(ELISA)的靈敏度相當,Spreeta SPR免疫傳感器的靈敏度有待提高。 (4)研究選用細胞裂解技術與Spreeta SPR免疫傳感器結合來降低檢測限,樣品菌液超聲處理后,檢測限下降為3×104 cfu mL-1,相關系數為0.98。結果表明細胞裂解技術能有效降低傳感器的檢測限。 (5)采用大腸桿菌DH 5α和李斯特菌作為驗證菌檢驗傳感器對大腸桿菌0157：H7的特異性,得出傳感器對目標菌的特異性良好,對驗證菌的吸附很小。
[Abstract]:E. coli 0157:H7 (E. coli 0157:H7, as one of the most harmful foodborne pathogens, has the possibility of epidemic outbreak and the severity of infection in China. Because of the disadvantages of complicated operation, laborious operation and time consuming in the traditional detection methods, it is urgent to develop a fast and accurate detection method. In this paper, many new detection techniques, including biosensors, have been investigated. According to the advantage of Spreeta SPR sensor, the Spreeta SPR sensor and cell lysis method were taken as the main research means and content, the detection of O157:H7 in E. coli was realized, and it was discussed and studied deeply. The main research contents and results are as follows: (1) the performance of Spreeta SPR sensor is evaluated with different concentrations of glucose solution. The correlation coefficient between glucose concentration and refractive index is close to 1, and the baseline is restored after rinsing with deionized water. The sensor has good performance. (2) the repeatability of the response results of avidin and biotinylated antibody showed that the sensor had high reproducibility, which provided a theoretical basis for the detection of Escherichia coli O157:H7 by Spreeta SPR immunosensor. The detection limit was 3 脳 10 ~ 6 cfu mL-1,. Basically meet the test needs. (3) comparing the sensitivity of BIACORE SPR immunosensor, the detection limit of direct detection of Escherichia coli O157:H7 is 3 脳 10 ~ 5 cfu mL-1,. The sensitivity of, Spreeta SPR immunosensor is comparable to that of (ELISA) by enzyme linked immunosorbent assay (Elisa). The sensitivity of, Spreeta SPR immunosensor needs to be improved. (4) the detection limit was decreased to 3 脳 10 ~ 4 cfu mL-1, after ultrasonic treatment with Spreeta SPR immunosensor and cell lysis technique. The correlation coefficient was 0.98. The results show that the cell lysis technique can effectively reduce the detection limit of the sensor. (5) Escherichia coli DH 5 偽 and Listeria were used to test the specificity of the biosensor to Escherichia coli 0157:H7. The results showed that the specificity of the biosensor to the target bacteria was good, and the adsorption of the validation bacteria was very small.
【學位授予單位】：浙江大學
【學位級別】：碩士
【學位授予年份】：2012
【分類號】：R155.5

【參考文獻】

相關期刊論文 前10條

1 王鋒;樊先平;王民權;;光學生物傳感器的研究進展[J];材料導報;2004年07期

2 張文浩;鄭先哲;初國超;;應用SPR生物傳感器檢測酸菜發(fā)酵液中大腸桿菌[J];東北農業(yè)大學學報;2010年06期

3 曹振新,粱大開,郭明江;光纖表面等離子體波傳感器的溫度特性的研究[J];光譜學與光譜分析;2003年01期

4 曾捷;梁大開;曾振武;杜燕;;基于SPR光譜分析的液體折射率測量研究[J];光譜學與光譜分析;2006年04期

5 唐倩倩;王劍平;蓋玲;葉尊忠;應義斌;;ATP生物發(fā)光法檢測E. coliO157∶H7的研究[J];光譜學與光譜分析;2009年02期

6 丁紅雷,毛旭虎,鄒全明,王豪舉;腸出血性大腸桿菌O157:H7的分子生物學檢測方法研究進展[J];國外醫(yī)學.臨床生物化學與檢驗學分冊;2005年08期

7 唐曉艷;張洪友;;SPR生物傳感器及其應用進展[J];中國畜牧獸醫(yī);2006年04期

8 葛晶,殷涌光,王凱;使用SPR生物傳感器快速檢測大腸桿菌E.Coli 0157∶H7[J];吉林大學學報(工學版);2005年02期

9 王燕琴;朱建勇;;腸出血性大腸桿菌O157:H7檢測方法研究進展[J];內蒙古農業(yè)科技;2010年03期

10 徐尚榮;;大腸桿菌O157∶H7的生物學特性[J];青海畜牧獸醫(yī)雜志;2008年04期

相關碩士學位論文 前2條

1 孫洋;腸出血性大腸桿菌0157多重PCR檢測及其stx基因缺失突變株的研究[D];中國人民解放軍軍需大學;2002年

2 陳思;腸出血性大腸桿菌O157：H7的PCR與免疫學檢測技術的研究[D];中國農業(yè)大學;2005年

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