生物標(biāo)志物傳感技術(shù)研究
發(fā)布時間：2019-02-22 17:37
【摘要】：慢性病是進(jìn)行性的、不能自然痊愈及很少能夠完全治愈的一類疾病,其所包含的疾病主要有糖尿病、高血脂、冠心病、腦卒中、高血壓、腫瘤等。慢性病已經(jīng)成為全世界幾乎所有國家成年人的最主要死因,而生物標(biāo)志物監(jiān)測是實現(xiàn)慢性病二級預(yù)防(早發(fā)現(xiàn)、早診斷、早治療)和對癥治療的基礎(chǔ),控制慢性病快速增長的基本途徑。 本論文主要研究多種慢性病相關(guān)生物標(biāo)志物快速檢測的方法和實現(xiàn)途徑,助力醫(yī)院臨床生物標(biāo)志物快速測試以及家庭自測,特別是糖尿病、高血脂、痛風(fēng)及腫瘤患者的自我監(jiān)測。研究工作主要包括以下幾個方面： (1)電化學(xué)酶傳感技術(shù)研究 解決了電化學(xué)酶傳感器的準(zhǔn)確性、一致性和穩(wěn)定性問題�；陔p電極測試系統(tǒng)中電子媒介體作用的新機制,選擇適當(dāng)?shù)臋z測電壓來減少干擾物發(fā)生電極反應(yīng)的概率。選用微反應(yīng)腔和快速水合的傳感層來縮短響應(yīng)時間,使得電子媒介體的反應(yīng)比干擾反應(yīng)優(yōu)先進(jìn)行,進(jìn)一步消除樣品中存在的其它電活性物質(zhì)的干擾。由于電子媒介體、雙電極檢測技術(shù)和短檢測時間等因素的協(xié)同作用,所制備的尿酸、葡萄糖、膽固醇生物傳感器均具有良好的抗干擾能力。正常生理性內(nèi)源物質(zhì)基本不干擾測定,外源性藥物除極端高值外亦不干擾。采用絲網(wǎng)印刷技術(shù)一步完成傳感試劑的固定,簡化了制備工藝,并且提高了傳感器間的一致性。通過優(yōu)化傳感層印漿組成,提高尿酸、葡萄糖、膽固醇生物傳感器的儲存穩(wěn)定性。所制備的尿酸傳感器的保存期為12個月以上,葡萄糖傳感器的保存期為18個月以上,膽固醇傳感器的保存期為10個月以上 (2)電化學(xué)免疫傳感技術(shù)研究 以腫瘤標(biāo)志物甲胎蛋白(AFP)為研究對象,解決了腫瘤標(biāo)志物電化學(xué)免疫傳感器研制中的四個共性問題：電化學(xué)標(biāo)記方法、電化學(xué)檢測方法、生物識別分子的固定化方法和免疫反應(yīng)模式選擇。利用電活性物質(zhì)甲酸二茂鐵標(biāo)記甲胎蛋白,通過免疫滲濾方式和競爭免疫反應(yīng),構(gòu)建了甲胎蛋白免疫滲濾型電化學(xué)傳感器。利用膠體金介導(dǎo)辣根過氧化物標(biāo)記抗體,通過免疫層析方式和夾心免疫反應(yīng),構(gòu)建了甲胎蛋白免疫層析型電化學(xué)傳感器。采用絲網(wǎng)印刷技術(shù)和微量非接觸點樣技術(shù)批量制備了上述兩種甲胎蛋白電化學(xué)免疫傳感器,相比于傳統(tǒng)免疫分析方法(如ELISA),兩種方法一致性很好。由于不需要頻繁的清洗步驟,該方法操作簡便,并且節(jié)省時間,適于診所和家庭自檢。 (3)條帶型半定量膠體金免疫層析傳感技術(shù)研究 解決了膠體金免疫層析傳感技術(shù)靈敏度低和不適于定量的問題,實現(xiàn)了三種前列腺癌腫瘤標(biāo)志物的半定量檢測。研究發(fā)現(xiàn)：單位面積硝酸纖維素膜上固定化的抗體越多,靈敏度越高；單位數(shù)量膠體金上標(biāo)記的抗體越少,靈敏度越高,但穩(wěn)定性越差。但是使用牛血清白蛋白來填補膠體金表面的空位和封閉膠體金表面未和抗體結(jié)合的位點,可以穩(wěn)定膠體金溶液和減少非特異性反應(yīng)。采用多測試線,所制備的快速膠體金免疫層析傳感器的檢測限和線性范圍均達(dá)到臨床的要求。密封于有干燥劑的塑料袋中常溫(25℃)保存,傳感器穩(wěn)定期大于6個月。臨床實驗研究表明,該方法與ELISA檢測結(jié)果符合率為97.6%,可以應(yīng)用于臨床,從而滿足了應(yīng)對前列腺癌普查篩選和預(yù)后監(jiān)測等快速檢測的需要。
[Abstract]:Chronic diseases are progressive, non-natural and rarely able to cure a class of diseases, including diabetes, hyperlipidemia, coronary heart disease, stroke, hypertension, tumor, etc. Chronic diseases have become the leading cause of death for almost all of the world's adults, and biomarker monitoring is the basis for secondary prevention of chronic diseases (early detection, early diagnosis, early treatment) and symptomatic treatment, and the basic way to control the rapid growth of chronic diseases. This paper mainly studies the method and the way of rapid detection of various chronic diseases related biomarkers, the rapid test of the clinical biomarker of the hospital and the self-supervision of the family self-test, in particular the diabetes, the hyperlipidemia, the gout and the tumor. The study mainly includes the following parties: Surface: (1) Electrochemical enzyme sensing The technical research has solved the accuracy, the consistency and the consistency of the electrochemical enzyme sensor. stability problem. Based on the new mechanism of the action of the electronic mediator in the double-electrode test system, the appropriate detection voltage is selected to reduce the occurrence electrode of the interference object. the reaction time is shortened by selecting a micro-reaction cavity and a rapidly-hydrated sensing layer, so that the reaction of the electronic mediator is preferentially carried out compared with the interference reaction, and the other electric activity existing in the sample is further eliminated. The prepared uric acid, glucose and cholesterol biosensor have good effects due to the synergistic effect of the electronic media, the double-electrode detection technology, the short detection time and the like. Anti-interference ability. The normal physiological endogenous substance does not interfere with the measurement, but the exogenous drug has extreme high value. The method of screen printing is adopted to complete the fixation of the sensing reagent, the preparation process is simplified, and the sensor is improved. the consistency between the uric acid, the glucose and the cholesterol biosensor is improved by optimizing the sensing layer printing paste composition, the storage life of the prepared uric acid sensor is more than 12 months, the storage period of the glucose sensor is more than 18 months, and the storage period of the cholesterol sensor is More than 10 months (2) electrochemical Immunosensor technology was used to study the tumor marker alpha-fetoprotein (AFP). The four common problems in the development of the electrochemical immunosensor for tumor markers are the electrochemical marking method, the electrochemical detection method and the immobilization method of the biological recognition molecule. and the alpha-fetoprotein is constructed by the immunofiltration method and the competitive immune reaction. A percolation type electrochemical sensor is prepared by using colloidal gold to mediate the horseradish peroxidase-labeled antibody, and using the immunochromatographic method and the sandwich immune reaction to construct the alpha-fetoprotein immunity. the electrochemical immunosensor of the two kinds of alpha-fetoprotein is prepared in batches by adopting a screen printing technology and a micro non-contact sample application technology, and compared with a traditional immunoassay method (such as an ELISA), the method has good consistency, and the method is simple and convenient to operate and save time, Suitable for clinic and home self-test. (3) Strip-type semi-quantitative glue The study of the immunochromatographic sensing technology of the body gold solves the problems of low sensitivity and not suitable for quantitative detection of the colloidal gold immunochromatographic sensing technology, and realizes three leading positions. It was found that the more the antibody immobilized on the nitrocellulose membrane in the unit area, the higher the sensitivity, the less the antibody labeled on the unit number of colloidal gold, The higher the sensitivity, the higher the stability. However, bovine serum albumin is used to fill the space on the surface of the colloidal gold and to close the site where the colloidal gold surface is not bound to the antibody, and the glue can be stabilized. bulk gold solution and reduced nonspecific reaction. The test limit of the fast colloidal gold immunochromatographic sensor prepared by using a multi-test line and is sealed in a plastic bag with a drying agent at normal temperature (25 DEG C). The clinical experiment shows that the coincidence rate between the method and the ELISA is 97.6%, which can be applied to the clinic, so as to meet the screening and screening of the prostate cancer.
【學(xué)位授予單位】：中南大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2012
【分類號】：R318.0
本文編號：2428446
[Abstract]:Chronic diseases are progressive, non-natural and rarely able to cure a class of diseases, including diabetes, hyperlipidemia, coronary heart disease, stroke, hypertension, tumor, etc. Chronic diseases have become the leading cause of death for almost all of the world's adults, and biomarker monitoring is the basis for secondary prevention of chronic diseases (early detection, early diagnosis, early treatment) and symptomatic treatment, and the basic way to control the rapid growth of chronic diseases. This paper mainly studies the method and the way of rapid detection of various chronic diseases related biomarkers, the rapid test of the clinical biomarker of the hospital and the self-supervision of the family self-test, in particular the diabetes, the hyperlipidemia, the gout and the tumor. The study mainly includes the following parties: Surface: (1) Electrochemical enzyme sensing The technical research has solved the accuracy, the consistency and the consistency of the electrochemical enzyme sensor. stability problem. Based on the new mechanism of the action of the electronic mediator in the double-electrode test system, the appropriate detection voltage is selected to reduce the occurrence electrode of the interference object. the reaction time is shortened by selecting a micro-reaction cavity and a rapidly-hydrated sensing layer, so that the reaction of the electronic mediator is preferentially carried out compared with the interference reaction, and the other electric activity existing in the sample is further eliminated. The prepared uric acid, glucose and cholesterol biosensor have good effects due to the synergistic effect of the electronic media, the double-electrode detection technology, the short detection time and the like. Anti-interference ability. The normal physiological endogenous substance does not interfere with the measurement, but the exogenous drug has extreme high value. The method of screen printing is adopted to complete the fixation of the sensing reagent, the preparation process is simplified, and the sensor is improved. the consistency between the uric acid, the glucose and the cholesterol biosensor is improved by optimizing the sensing layer printing paste composition, the storage life of the prepared uric acid sensor is more than 12 months, the storage period of the glucose sensor is more than 18 months, and the storage period of the cholesterol sensor is More than 10 months (2) electrochemical Immunosensor technology was used to study the tumor marker alpha-fetoprotein (AFP). The four common problems in the development of the electrochemical immunosensor for tumor markers are the electrochemical marking method, the electrochemical detection method and the immobilization method of the biological recognition molecule. and the alpha-fetoprotein is constructed by the immunofiltration method and the competitive immune reaction. A percolation type electrochemical sensor is prepared by using colloidal gold to mediate the horseradish peroxidase-labeled antibody, and using the immunochromatographic method and the sandwich immune reaction to construct the alpha-fetoprotein immunity. the electrochemical immunosensor of the two kinds of alpha-fetoprotein is prepared in batches by adopting a screen printing technology and a micro non-contact sample application technology, and compared with a traditional immunoassay method (such as an ELISA), the method has good consistency, and the method is simple and convenient to operate and save time, Suitable for clinic and home self-test. (3) Strip-type semi-quantitative glue The study of the immunochromatographic sensing technology of the body gold solves the problems of low sensitivity and not suitable for quantitative detection of the colloidal gold immunochromatographic sensing technology, and realizes three leading positions. It was found that the more the antibody immobilized on the nitrocellulose membrane in the unit area, the higher the sensitivity, the less the antibody labeled on the unit number of colloidal gold, The higher the sensitivity, the higher the stability. However, bovine serum albumin is used to fill the space on the surface of the colloidal gold and to close the site where the colloidal gold surface is not bound to the antibody, and the glue can be stabilized. bulk gold solution and reduced nonspecific reaction. The test limit of the fast colloidal gold immunochromatographic sensor prepared by using a multi-test line and is sealed in a plastic bag with a drying agent at normal temperature (25 DEG C). The clinical experiment shows that the coincidence rate between the method and the ELISA is 97.6%, which can be applied to the clinic, so as to meet the screening and screening of the prostate cancer.
【學(xué)位授予單位】：中南大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2012
【分類號】：R318.0
【參考文獻(xiàn)】
相關(guān)期刊論文 前6條
1 杜冬梅;任穎;魏盟;邵琦;趙催春;;空腹血糖水平對初發(fā)急性心肌梗死患者預(yù)后的影響[J];上海交通大學(xué)學(xué)報(醫(yī)學(xué)版);2010年04期
2 劉麗強;彭池芳;金征宇;胥傳來;;納米金技術(shù)的發(fā)展及在食品安全快速檢測中的應(yīng)用[J];食品科學(xué);2007年05期
3 龐文艷;袁如玉;葉嵐;黃嬌紅;李立豐;李廣平;;高密度脂蛋白膽固醇與冠心病及其危險因素的回顧性分析[J];天津醫(yī)科大學(xué)學(xué)報;2009年02期
4 楊自華,何林,周克元;檢驗醫(yī)學(xué)干化學(xué)分析技術(shù)進(jìn)展[J];現(xiàn)代儀器;2005年04期
5 肖紅玉;柴春彥;劉國艷;史賢明;;檢測動物性食品中克倫特羅殘留的免疫電化學(xué)傳感器反應(yīng)體系的確定[J];中國食品學(xué)報;2006年01期
6 馬靜;張偉尉;李聞;陳曉春;劉紅艷;黃正;;基于納米金固定大腸桿菌O157:H7酶免疫傳感器的研究[J];中國衛(wèi)生檢驗雜志;2007年12期
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