Cloning, Expression, Purification of NSE and Preparation of NSE Polyclonal Antibody

神经元特异性烯醇化酶(Neuronspecificenolase)是糖酵解途径中一种重要的生物酶，其特异性存在于神经元、神经内分泌细胞中。血清NSE水平的变化作为敏感的病理学标志蛋白以及中枢神经系统损伤时的定量指标正逐渐被重视，且血清NSE已成为小细胞肺癌(SCLC)诊断中最敏感的肿瘤标志物。随着研究的进一步深入，非小细胞肺癌(NSCLC)神经内分泌化逐步引起人们的重视，神经内分泌产物NSE在NSCLC中的地位也越来越受关注。总之，NSE作为神经内分泌性质肿瘤的标志物及神经元损伤的标志物，已越来越受国内外学者的关注。 在这个大背景下，为了进一步研究NSE在肿瘤与临床方面的意义与应用。我们通过...Neuron specific enolase (NSE) that specifically exists in neurons and neuroendocrine cells is a key enzyme in glycolysis pathways. NSE as a sensitive protein marker in pathology and a quantitative index of central nervous system damage is gaining more and more attention. Serous NSE has been considered as the most senstive tumor marker in diagnosis of SCLC. Along with the further studies, the neuro...学位：理学硕士院系专业：生命科学学院生物学系_生物化学与分子生物学学号：2172008115264

Characterization of NSE monoclonal antibodies and establishment of a double-antibody sandwich ELISA assay

通讯作者，E-mail: xtli@ xmu.edu.cn 作者简介: 丁焕弟( 1985 年－ ) ，女，在读硕士，主要从事抗肿瘤单克 隆抗体及诊断试剂盒的研究，E-mail: dinghuandi1125 @163.com。[中文文摘]目的:制备并鉴定NSE(Neuron-specific enolase)单克隆抗体,建立可检测NSE蛋白的双抗夹心ELISA方法。方法:用本实验室已表达纯化的NSE融合蛋白免疫BALB/c小鼠,采用杂交瘤技术制备单克隆抗体。采用WB、IP、IF、IHC等方法对获得的NSE单抗进行鉴定及亚型检测。利用辣根过氧化物酶标记纯化后的NSE单抗,建立一个可检测NSE蛋白的双抗夹心ELISA法。结果:通过分析和鉴定,选定2株可稳定分泌抗NSE抗体的杂交瘤细胞株,效价达4.2×107～6.5×107,亚型为IgG2b。免疫印迹结果显示,该抗体不仅能识别细胞内源NSE蛋白,还能识别分泌到细胞培养上清液中的NSE蛋白,此外还可用于免疫荧光及免疫组化检测。文中所建立的双抗夹心ELISA法,最低检测极限为8.85 ng/ml。结论:成功获得了效价高、灵敏度好及特异性强的NSE单抗,建立了一个双抗体夹心ELISA检测系统,具有良好的临床应用前景。[英文文摘]Objective: Preparation and characterization of monoclonal antibodies against NSE protein，and establishment of a double-antibody sandwich ELISA assay． Methods: BALB/c mice were immunized by using purified recombinant NSE，and monoclonal antibodies were generated by hydridoma technique． These antibodies were characterized with ELISA，Western blot，Immunofluorescent and Immunohistochemical staining． The isotypes of these antibodies were determined with an antibody isotyping kit． With Horseradish Peroxidase labelled NSE monoclonal antibody，we were able to establish a double-antibody sandwich ELISA to detect NSE protein． Results: Two positive hybridoma cell lines were selected for test，the titers of these two monoclonal antibodies could reach 4． 2 × 107 －6. 5 × 107，and their isotypes were IgG2b． Our NSE antibodies could detect not only endogernous NSE protein from cells，but also secreted NSE protein from cells in culture medium by Western blot，in addition，they could be used for immunofluorescent and immunohistochemical staining． The minimum amount of NSE protein could be detected by this double-antibody sandwich ELISA was 8． 85 ng /ml． Conclusion: Our NSE monoclonal antibodies achieved good sensitivity and specificity with high titers，and we established a doubleantibody sandwich ELISA assay which could be used for clinical test in future．福建省科技重点项目(编号项目No.2011Y0050); 厦门市科技计划项目(No.3502Z20123009

シンシッカン ニ オケル ウンドウ リョウホウ ノ ユウヨウセイ

Cardiac rehabilitation has recently become recognized as a safe treatment to care patients withcardiovascular diseases. To date, cardiac rehabilitation program has been established by various clinical studiesand statements. Cardiac rehabilitation prescribes exercise that has to be performed under the level of lactic acidproduction. Individualized exercise program serves as a secondary prevention of ischemic heart disease,improves cardiac function, and possesses an effect of decreasing an incidence of ventricular arrhythmia. Thus,cardiac rehabilitation that can apply to most of patients with cardiovascular diseases is expected to be widelyendorsed in order to reduce cardiovascular mortality and morbidity and optimize health in persons withcardiovascular diseases

コロナ禍地方急性期病院におけるロボット支援下手術の導入

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思春期における悪性腫瘍患者の看護 その2 : 積極的に登校を試みた一症例を通して

Article信州大学医学部附属病院看護研究集録 1985: 27-36 (1985)departmental bulletin pape