低分子量紫色酸性磷酸酶多克隆抗体的制备及应用

将最近从地瓜中提出来的低分子量紫色酸性磷酸酶（smPAP)基因克隆到GST融合蛋白表达载体pGEX－2T中，在大肠杆菌BL21codon plus中进行表达，用表达的融合蛋白免疫兔子产生多克隆抗体，用抗血清在昆虫细胞中表达的smPAP和地瓜提取液中分离纯化的PAP进行检测，产生了良好的交叉反应。     

hBAF53 多克隆抗体的制备与活性分析

用PCR方法扩增了人的BAF53全长eDNA序列，用酶切后的△BAF53 eDNA片段构建了表达质粒．转化大肠杆菌后，经IPTG诱导表达，再经包涵体裂解、纯化，得到hBAF53抗原多肽．从免疫动物身上获得实验动物的抗血清．通过斑点印迹方法测得hBAF53抗血清的特异性和效价，又用已提取的HeLa细胞核蛋白(含有天然BAF53蛋白)进行免疫印迹分析，证明天然。BAF53蛋白是该抗血清的抗原，说明获得的抗血清具有比较高的特异性，可用于染色体改构复合物各亚基问相互作用分析及调控基因转录机理方面的研究．     

反相液相色谱法同时检测虾仁中合成色素的研究

建立了食品虾仁中合成色素(酸性大红GR、金黄粉、橙黄G、酸性红1号、酸性红26)的反相液相色谱同时检测方法。并且对提取剂、NH4Ac浓度和离子对种类等因素进行优化,使得5种色素完全分离,并且在色谱图上能与国标内的6种色素完全分离。金黄粉、酸性大红GR、橙黄G、酸性红1号、酸性红26的回收率均在80%以上,在0.05~10μg/mL范围内有良好的线性关系,相关系数大于0.9998。本法可用于食品虾仁中合成着色剂的快速检测。     

抗酮洛芬多克隆抗体的制备及其间接竞争ELISA检测方法的建立

分别将酮洛芬与牛血清白蛋白(BSA)及卵清蛋白(OVA)偶联制得免疫原和包被原,经过免疫新西兰白兔制备多克隆抗体,抗体经纯化后效价为1:128000。使用自制的抗体,建立了测定酮洛芬的间接竞争酶联免疫吸附(ic-ELISA)新方法。ic-ELISA的线性范围为0.010～10.0μg/L,IC50为0.235μg/L,最低检测限为0.0040μg/L,线性回归方程为y=-22.97ρ+104.5(R2=0.980),与布洛芬、双氯酚酸的交叉反应率均小于4%,方法可用于水体中酮洛芬的检测。     

玉米赤霉醇人工抗原合成及其多克隆抗体的制备

对玉米赤霉醇(ZER)16-OH进行改造,设计合成了模拟玉米赤霉醇特征结构的半抗原———ZER-16-羧丙基丁醚。用混合酸酐法将其与载体蛋白BSA连接,用于免疫新西兰大白兔,制备的抗血清效价达1∶102 400。基于活性酯法合成的包被抗原建立了CI-ELISA检测方法,该法对玉米赤霉醇检测的线性范围为18.39~1744.70 ng/mL,检出限为8.61 ng/mL。     

毕兹多克隆抗体的制备和分析应用

用琥珀酸酐－碳二亚胺法和氯磺化法合成了毕兹（Ｂｚ）的两种人工抗原Ｂｚ＿ＨＳ＿ｐｒｏｔｅｉｎ和Ｂｚ＿ＳＯ２＿ｐｒｏｔｅｉｎ。由Ｂｚ＿ＳＯ２＿ＢＳＡ（牛血清白蛋白）诱导出的抗血清能够用来测定１０－９ｍｏｌ·Ｌ－１的Ｂｚ，工作曲线的线性范围在３×１０－５～３×１０－９ｍｏｌ·Ｌ－１，相对标准偏差为１３６％（ｎ＝６），回收率在９０％～１１５％。     

酸性大红3R的电化学研究及应用

采用循环伏安法和方波伏安法考察了酸性大红3R在碳糊电极上的伏安行为和反应机理。当扫描速率为0.1 V/s时,在pH 1的0.05 mol/L H2SO4支持电解液中,酸性大红3R有一对峰电位分别是0.74 V和0.72 V（vs.饱和甘汞电极）的氧化峰和还原峰,并且显示吸附控制的2电子单质子的准可逆氧化还原过程。在最佳实验条件下,酸性大红3R的方波氧化峰峰电流与其浓度在2.12×10^-5-3.39×10^-4mol/L范围内呈线性关系,检出限为8.35×10^-6mol/L,由此建立测定酸性大红3R含量的方波伏安法。该方法可用于计算酸性大红3R常规染羊毛与蚕丝的上染率,测定结果与分光光度法一致。     

酸性媒介红稀土染料的制备、表征及其羊毛染色性能

制备并提纯了酸性媒介红稀土(La, Nd, Y, Er)染料, 稀土及元素分析测得其稀土与配体酸性媒介红染料物质的量的比为1∶1; 红外光谱、紫外-可见光谱分析表明, 在酸性媒介红稀土染料的结构中, 稀土离子(La3+, Nd3+, Y3+)与配体染料分子中的羧基、羟基产生配位作用, 其中Y3+同时还与偶氮基(-N=N-)作用,在3个稀土染料中稀土离子与染料分子中的磺酸基(-SO3-)也产生作用; 所有样品的羊毛布料染色实验及其各项牢度测试结果表明 酸性媒介红稀土染料羊毛染色比相应配体酸性媒介红染料色光纯正, La, Nd染料染色增深, Y, Er染料染色增艳, 但均比铬金属络合染料颜色鲜艳, 钇染料的各项羊毛染色牢度比配体染料好, 与Cr金属络合染料相当, 具有实用价值.     

利用多克隆抗体有效识别中草药中抗表皮生长因子抑制剂

用一种已知的抗表皮生长因子受体抑制剂 (piceatannol) 作为半抗原与载体牛血清白蛋白 (BSA) 连接后免疫制备相应的多克隆抗体 (PcAb).利用该多克隆抗体来模拟酶制成亲和色谱柱，从一种藏药粗提物中将包括该半抗原在内的几种结构不同的抗表皮生长因子受体抑制剂识别出来.研究采用前沿亲和色谱-质谱联用技术对样品进行分析，可以直接从中药复杂体系中识别出有效成分并进行鉴定，实现中药有效成分的筛选与鉴定一体化技术.     

酸性红褪色分光光度法测定食盐中碘的含量

研究了在H_2SO_4-KBr的介质中,碘酸钾能够氧化酸性红并使之褪色,且褪色的程度与碘酸根的含量在一定范围内有很好的线性关系,从而建立了一种新的测定碘的方法。实验结果表明,酸性红的最大吸收波长为510nm,碘的含量在0～50μg/50mL范围内,符合比耳定律,其回归方程为△A=0.004 13C+0.032 14,相关系数R2=0.998 3,检出限为0.78μg/50mL。方法操作简单,快速准确,用于食盐中碘含量的测定,结果令人满意。     

Prokaryotic Expression and Polyclonal Antibody Preparation of PRL3

Phosphatase of regenerating liver 3(PRL3),which belongs to the superfamily of protein tyrosine phosphatases(PTPs),represents a group of low molecular weight PTPs that participate in tumorigenesis and metastasis processes.Presented here are the results of cloning,prokaryotic expression,purification,and polyclonal antibody preparation of PRL3.To obtain a specific polyclonal antibody against PRL3,the authors have prepared GST-PRL3 to immunize rabbits and purify an anti-PRL3 polyclonal antibody by negative selection affinity columns.Western blot analysis shows that the anti-PRL3 polyclonal antibody has a specific binding ability with PRL3 protein.The anti-PRL3 polyclonal antibody provides a good tool to further study the function of PRL3.     

Construction and Expression of Human Survivin and Preparation of Its Polyclonal Antibody

Survivin, a novel member of inhibitor of apoptosis（IAP） protein family, is aberrantly expressed in cancer but undetectable in normal, differentiated adult tissues. The cancer-specific expression of survivin, coupled with its importance in inhibiting cell death and in regulating cell division makes it a useful diagnostic marker of cancer and a potential target for cancer treatment. Survivin cDNA amplified from the total RNA of 293 cells through RT-PCR was cloned into prokaryotic expression vector pRSET-B. The recombinant plasmid pRSET-B-Surv was expressed in E.coli BL21, and the relative molecule mass（Mr） of expressed fusion protein was approximately 21000. The recombinant protein was purified through Ni^2＋ affinity chromatography column and characterized by SDS-PAGE and Western blot. The purified recombinant protein was then injected into rabbits, and antisurvivin polyclonal antibody with a high titer was obtained.     

Preparation of Polyclonal Antibodies Against Testis-specific Protease 50 and Characterization of Antibody Specificity

Introduction TheTSP50genehasbeenidentifiedasoneofthe testis specificoncogenes,whichisexpressedathigh levelsinapproximately92%ofhumanbreastcancer samples,makingitanattractivemolecularmarkerand apotentialtargetfordiagnosisandtherapy[1].Itisho mologoustomany…     

Expression of Catalytic Domain of Protein Tyrosine Phosphatase 1B and Preparation of Its Polyclonal Antibody

This study focuses on the expression of human protein tyrosine phosphatase 1B(PTP1B) catalytic domain (△PTP1B) and preparation of polyclonal antibody against △PTP1B. △PTP1B gene was PCR amplified with the cDNA of human PTP1B as the template, and cloned into the pT7 expression vector. The recombinant pT7-△PTP1B was expressed in E. coli Rosetta( DE3 ) host cells and purified. The antiserum was prepared by immunizing rabbit with purified recombinant △PTP1B. The polyclonal antibody against △PTP1B was purified by PVDF immobilized antigen affinity chromatography. △PTP1B was correctly cloned, expressed, and purified as confirmed by PCR, DNA sequence ratio) and 0. 1 ng, respectively. This study provides an important basis for further studying the biological function of PTP1B and its relationship with human diseases.     

GST Fusion Protein Based Specific Polyclonal Antibody Preparation of Mouse Aquaporin 1

Aquaporins(AQPs) are specific membrane channels for water and other small nonionic molecules.In order to overcome the difficulties to generate the effictive antibody of membrane protein,we selected the cytoplasmic C-terminus of Aquaporin 1(AQP1) as an unique antigen.The long C-terminus of mouse AQP1 was overexpressed in the Glutathione S-tansferase Gene Fusion System.On the basis of the resonable amounts of soluable membrane protein peptides,we prepared the specific antibody.To pursure this object,we constr...     

Preparation and Characterization of Olaquindox Polyclonal Antibody

本研究先将喹乙醇琥珀酸单酯化,使其转变为带有羧基的衍生物。合成产物经重结晶纯化,得率为52.47%。合成产物进行结构鉴定,合成了具有喹乙醇分子结构特征的喹乙醇半抗原。采用活化酯法将半抗原与牛血清白蛋白（BSA）进行偶联,采用混合酸酐法将半抗原与卵清蛋白（OVA）进行偶联,分别制备免疫原和包被抗原。紫外扫描与红外图谱分析结果表明,半抗原与载体蛋白偶联成功,与BSA、OVA的结合比分别为3.8:1和5.0:1。以OLA-BSA作为免疫原免疫4只新西兰大白兔,获得了较高效价的抗血清,以OLA-OVA作为包被抗原,间接ELISA法测定各兔的效价,分别为1:6400;1:1600;1:12800;1:6400。间接ELISA法喹乙醇的抑制中浓度IC50为743.3 ng/mL,最低检测限IC20为5.71 ng/mL。从电泳图谱分析可以看出,经纯化得到了纯度较高的喹乙醇多克隆抗体,为喹乙醇免疫分析方法的进一步研制和开发奠定基础。     

稀土上转换发光材料标记抗体的制备及在免疫组化中的应用

采用微波辐射法合成了具有上转换发光特性的六方相纳米粒子NaGdF₄: Yb³⁺,Er³⁺(UCNPs), 其晶粒大小约为65 nm, 且粒子在980 nm的激发光下显示绿光(550 nm). 进一步在NaGdF₄: Yb³⁺,Er³⁺纳米晶的表面包覆了一层二氧化硅层, 进行氨基功能化后获得了表面共价结合氨基基团的粒径为70 nm的上转换发光纳米微球NaGdF₄: Yb³⁺,Er³⁺@SiO₂-NH₂(UCNPs@SiO₂-NH₂). 通过共价键将UCNPs@SiO₂-NH₂与多克隆抗体免疫球蛋白联接, 将标记后的多克隆抗体应用于传统的免疫组化检测子宫内膜腺细胞中基质金属蛋白酶组织抑制剂-4(TIMP-4)蛋白的表达. 结果表明, 微波合成的稀土上转换发光纳米材料形貌规则且粒径均一, 包覆硅壳后材料具有良好的分散性和水溶性, 荧光强度高且稳定, 在980 nm激发光下对生物组织无背景荧光, 可以很好地检测组织中蛋白质的表达.