本文選題：鉛　切入點(diǎn)：鎘　出處：《吉林大學(xué)》2011年碩士論文

【摘要】：本研究通過雙功能螯合劑將鉛、鎘離子分別與牛血清白蛋白(BSA)和卵清白蛋白(OVA)偶聯(lián),合成免疫抗原Pb-ITCBE-BSA、Cd-ITCBE-BSA和檢測抗原Pb-ITCBE-OVA、Cd-ITCBE-OVA,采用SDS-PAGE電泳法和紫外分光光度法對制備的抗原進(jìn)行鑒定。足墊注射方法免疫Balb/c小鼠,間接ELISA方法檢測小鼠血清效價,采用PEG方法進(jìn)行細(xì)胞融合,間接ELISA方法篩選陽性雜交瘤細(xì)胞,有限稀釋法克隆雜交瘤細(xì)胞株,獲得了抗鉛離子的單克隆抗體細(xì)胞株3C1和抗鎘單克隆抗體細(xì)胞株5E12。兩株單抗細(xì)胞株的亞類均為IgG1,親和力常數(shù)分別為1.19×108和1.02×108,其他金屬離子的交叉反應(yīng)均小于3%。通過體內(nèi)誘生法制備腹水型單抗,采用親和層析法純化腹水。分別用3C1和5E12純化的腹水建立優(yōu)化了ELISA方法。采用間接競爭ELISA方法檢測鉛、鎘標(biāo)準(zhǔn)品,并建立標(biāo)準(zhǔn)曲線,其中抗鉛單克隆抗體的標(biāo)準(zhǔn)曲線回歸方程是y=y=-16.325x+114.53,相關(guān)系數(shù)R2=0.9873,檢測范圍是20-160ng/mL,最低檢測限為20ng/mL;抗鎘單克隆抗體的標(biāo)準(zhǔn)曲線方程為y=-13.345+103.42,回歸系數(shù)R2=0.994,檢測范圍是0.01-0.64μg/mL,最低檢測限為0.01μg/mL。分別在純凈水、自來水和南湖水樣中添加鉛、鎘標(biāo)準(zhǔn)品溶液,添加回收率分別為99.73%、101.01%、96.83%和101.62%、95.75%、98.49%。本研究為鉛、鎘離子酶聯(lián)免疫快速檢測試劑盒和免疫膠體金試紙條的開發(fā)利用奠定了一定的基礎(chǔ)。
[Abstract]:In this study, lead and cadmium ions were coupled with bovine serum albumin (BSA) and ovalbumin (OVA) respectively to synthesize immune antigen Pb-ITCBE-BSA-Cd-ITCBE-BSA and detection antigen Pb-ITCBE-OVACd-ITCBE-OVA. the prepared antigens were identified by SDS-PAGE electrophoresis and UV spectrophotometry.Balb/c mice were immunized with foot pad injection method, serum titers of mice were detected by indirect ELISA method, cell fusion was performed by PEG method, positive hybridoma cells were screened by indirect ELISA method, hybridoma cell lines were cloned by limited dilution method.The monoclonal antibody cell line 3C1 and the monoclonal antibody cell line 5E12 were obtained.The affinity constants of the two McAb cell lines were 1.19 脳 10 ~ 8 and 1.02 脳 10 ~ 8, respectively. The cross reaction of other metal ions was less than 3.Ascites were purified by affinity chromatography.ELISA method was established by using 3C1 and 5E12 purified ascites.Indirect competitive ELISA method was used to detect lead and cadmium standard, and the standard curve was established.Lead and cadmium were added to pure water, tap water and Nanhu water, respectively. The recovery rates were 99.73 and 101.01% and 101.62% and 95.75%, respectively.This study laid a foundation for the development and utilization of the rapid detection kit for lead and cadmium ions and the immunocolloidal gold strip.
【學(xué)位授予單位】：吉林大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2011
【分類號】：R392

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