本文選題：FL + 雜交瘤��； 參考：《蘇州大學(xué)》2010年碩士論文

【摘要】： 目的 研制鼠抗人Flt3-ligand(FL)單克隆抗體(mAb)并對其生物學(xué)功能進行研究。 方法 采用高表達膜FL的基因工程細(xì)胞株L929/FL細(xì)胞免疫BALB/c小鼠,利用B淋巴細(xì)胞雜交瘤技術(shù)進行細(xì)胞融合,免疫熒光標(biāo)記和流式細(xì)胞術(shù)篩選陽性株,采用快速定性試紙法鑒定單抗亞類、間接免疫熒光法檢測單抗對腫瘤細(xì)胞株的識別譜,然后研究所獲得的抗人FL單克隆抗體及FL蛋白對腫瘤細(xì)胞生長的影響。 結(jié)果 得到穩(wěn)定分泌鼠抗人FL單克隆抗體的雜交瘤細(xì)胞三株,命名為5H5、5D1、3E5,快速定性試紙法鑒定5H5、5D1屬小鼠IgM亞類及κ型,3E5屬小鼠IgG1亞類及κ型;經(jīng)體內(nèi)誘生腹水法制備抗體,小鼠腹水形成的陽性率約為80%,腹水的收獲量平均為2 ml/只小鼠;經(jīng)免疫親和層析法純化,腹水中抗體蛋白的得率為1.0 mg/ml;用流式細(xì)胞術(shù)分析細(xì)胞膜熒光標(biāo)記百分率和熒光強度,結(jié)果顯示,FL能不同程度地在某些腫瘤細(xì)胞株上表達。將FL蛋白和3E5加入到U937、675321、THP-1細(xì)胞的培養(yǎng)體系中,經(jīng)細(xì)胞計數(shù)分析,3E5能夠阻斷FL蛋白對U937、675321、THP-1細(xì)胞生長的促進作用。 結(jié)論 本研究獲取三株分泌鼠抗人FL單克隆抗體的雜交瘤細(xì)胞,且3E5為一株阻斷型單克隆抗體,這為研究FL及Flt3功能奠定了物質(zhì)基礎(chǔ)。
[Abstract]:Purpose Mouse anti-human Flt 3 ligand FLL) monoclonal antibody was prepared and its biological function was studied. Method BALB/c mice were immunized with genetically engineered cell line (L929/FL) with high expression membrane FL. The BALB/c mice were fused with B lymphocyte hybridoma technique. The positive strains were screened by immunofluorescence labeling and flow cytometry. The monoclonal subclasses were identified by rapid qualitative test paper method. Indirect immunofluorescence assay was used to detect the recognition spectrum of the monoclonal antibody against human FL and the effect of FL protein on the growth of tumor cells. Result Three hybridoma cells stably secreting monoclonal antibody against human FL were obtained, named as 5H5H5O5D1O3E5. Rapid qualitative test paper was used to identify the subclasses of IgM and IgG1 and 魏 type of 5H5D5D1D1E5 and 5H5D5E5, respectively, and to prepare the antibody by the method of inducing ascites in vivo. The positive rate of ascites formation in mice was about 80%, the average yield of ascites was 2 ml/ in mice, the yield of antibody protein in ascites was 1.0 mg / ml by immunoaffinity chromatography, and the fluorescence labeling percentage and fluorescence intensity of cell membrane were analyzed by flow cytometry. The results showed that FL could be expressed in some tumor cell lines to some extent. FL-protein and 3E5 were added to the culture system of U937O675321 THP-1 cells. The cell count analysis showed that FL-protein could block the growth of U937C675321 cells. Conclusion In this study, three hybridoma cells secreting mouse anti-human FL monoclonal antibody were obtained, and 3E5 was a blocking monoclonal antibody, which laid a material foundation for studying the function of FL and Flt3.
【學(xué)位授予單位】：蘇州大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2010
【分類號】：R392

【共引文獻】

相關(guān)博士學(xué)位論文 前1條

1 何永林;GLS/IL-12重組沙門菌抗腫瘤的實驗研究[D];重慶醫(yī)科大學(xué);2007年

，

本文編號：1942227