【摘要】： 本課題以長白白眉蝮蛇蛇毒為原料，通過DEAE-Sephadex A-50離子交換層析、Sephadex G-75凝膠層析、Lysine Sepharose TM4B層析得到蛇毒纖溶酶原激活劑。以此為抗原對(duì)BALB／C小鼠進(jìn)行免疫，通過雜交瘤技術(shù)獲得免疫BALB／C小鼠脾細(xì)胞和小鼠骨髓瘤細(xì)胞的陽性融合細(xì)胞株，使用定量的陽性融合細(xì)胞誘導(dǎo)小鼠產(chǎn)生效價(jià)高的腹水，將小鼠腹水用辛酸．硫酸銨法進(jìn)行一系列的純化。共獲得4株長白白眉蝮蛇蛇毒纖溶酶原激活劑單克隆抗體，為287、3C11、4E7、4H10。進(jìn)行較系統(tǒng)的免疫特性的鑒定：4株單抗特異性高，與u-PA、BSA均無交叉反應(yīng)；腹水效價(jià)8200～10000；免疫球蛋白亞類為IgG1和IgG2a；4株抗體中，287和3C11可明顯抑制長白白眉蝮蛇蛇毒纖溶酶原激活劑活性，而4E7、4H10則對(duì)其活性無明顯影響。287和3C11這2株抗體為建立ELISA法檢測長白白眉蝮蛇蛇毒纖溶酶原激活劑含量以及研究其功能和結(jié)構(gòu)的關(guān)系奠定了基礎(chǔ)，，并為該成分的親和層析提供重要配體。
[Abstract]:In this study, the venom plasminogen activator of snake venom was obtained by DEAE-Sephadex A-50 ion exchange chromatography Sephadex G-75 gel chromatography and Lysine Sepharose TM4B chromatography using Agkistrodon halys venom as raw material. The positive fusion cell lines of spleen cells and myeloma cells of BALB/C mice were obtained by hybridoma technique. The positive fusion cells were induced to produce ascites with high titer by using quantitative positive fusion cells. Mouse ascites were purified by octanoic acid and ammonium sulfate method. Four monoclonal antibodies against plasminogen activator of Agkistrodon acutus venom were obtained. The specific of the McAbs was high, and there was no cross reaction with u-PABSA. The ascites titer was 8200,10000. the immunoglobulin subclasses were IgG1 and IgG2a;. The activity of plasminogen activator in venom of Agkistrodon halys was significantly inhibited by 3C11. However, 4E7H _ (10) had no significant effect on the activity of. 287 and 3C11, which laid a foundation for the detection of plasminogen activator in Agkistrodon halys venom by ELISA, and for the study of the relationship between the function and structure of plasminogen activator of Agkistrodon halys venom. It also provides an important ligand for affinity chromatography.
【學(xué)位授予單位】：沈陽藥科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2005
【分類號(hào)】：R392

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