制备和鉴定脑靶向性脂质体P-MMA-DOSPER

发布时间：2018-04-01 14:25

本文选题：侧脑室　切入点：脂质体　出处：《第一军医大学》2006年硕士论文

【摘要】：研究背景：中枢神经系统基因治疗的穿梭载体主要有病毒载体和非病毒载体两类，由于两者有着各自不同的缺点，还不能作为理想的基因载体运用于临床。而载体的选择是基因治疗的关键，所以研究者们不遗余力的寻找安全、高效、稳定的基因载体。相对于病毒载体脂质体优点在于安全、无免疫源性，缺点是基因转移效率低。因此，脂质体的研究主要集中于提高基因转移效率方面。偶联了抗体的免疫脂质体就是通过抗体的靶向特异性实现脂质体在某些组织器官的聚集来提高基因转移效率。目的 1．探求将脂质体构建成为更理想的脑内基因转移穿梭载体的方法。2．以神经干细胞表面抗原为靶位，以衍生化单克隆抗体偶联脂质体脑内寻靶，使基因治疗更具目的性，提高基因转移效率。方法 1．使用LB(Luria-Bertani)培养基扩增培养pEGFP-C_2质粒转化后的JM—109大肠杆菌，以质粒抽提试剂盒提取大量pEGFP-C_2质粒。经过酶切电泳、DNA测序鉴定并以紫外分光光度法检测质粒浓度及纯度后用于基因转染。2．以二棕榈酸磷脂酰乙醇胺(DPPE)对单克隆抗体(anti-Lex／SSEA-1)进行衍生化，采用透析法分离纯化衍生化抗体、紫外分光光度法检测抗体衍生化的效率，，间接荧光抗体实验法检验衍生化后抗体的活性。实验中用碳二亚胺法把衍生化的单克隆抗体P-MMA与阳离子脂质体DOSPER共价偶联，制成有脑靶向性的
[Abstract]:Background: central nervous system gene therapy of shuttle vector mainly viral and non viral vectors in two categories, because of their respective shortcomings, also cannot be used as an ideal gene vector for clinical use. The vector selection is the key of gene therapy, so the researchers spare no effort to find a safe, efficient, gene the carrier is stable. Compared with viral vector liposome has the advantages of safety, no immunogenicity, disadvantage is the low gene transfer efficiency. Therefore, the research of liposome mainly focused on improving the gene transfer efficiency. The coupling of immune liposome by antibodies targeting specific implementation of aggregation of liposomes in some organs. To improve the efficiency of gene transfer.
Objective 1. to explore the way to construct liposome into an ideal shuttle vector for brain gene transfer..2. targets the neural stem cell surface antigen, and targets in the brain with monoclonal antibody coupled with liposomes. It makes gene therapy more purposeful and improves gene transfer efficiency.
1. methods of using LB (Luria-Bertani) medium cultured pEGFP-C_2 plasmid after JM 109 Escherichia coli, extracted from a large number of pEGFP-C_2 plasmid to Plasmid Extraction Kit. After enzyme digestion, DNA sequencing and the UV spectrophotometric detection of plasmid concentration and purity with.2. in gene transfection with two palmitic acid phospholipid phosphatidylethanolamine (DPPE) of monoclonal antibodies (anti-Lex / SSEA-1) for derivatization, derivatization of separation and purification of antibodies by dialysis, derivatization efficiency of UV spectrophotometric detection of antibody, indirect fluorescent antibody test antibody activity test after derivatization. Experiments using two carbon imine derivatization method monoclonal antibody P-MMA and cationic liposome DOSPER covalent coupling, made of brain targeting

【学位授予单位】：第一军医大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R346

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