發(fā)布時(shí)間：2018-04-01 14:25

  本文選題：側(cè)腦室　切入點(diǎn)：脂質(zhì)體　出處：《第一軍醫(yī)大學(xué)》2006年碩士論文

【摘要】：研究背景：中樞神經(jīng)系統(tǒng)基因治療的穿梭載體主要有病毒載體和非病毒載體兩類，由于兩者有著各自不同的缺點(diǎn)，還不能作為理想的基因載體運(yùn)用于臨床。而載體的選擇是基因治療的關(guān)鍵，所以研究者們不遺余力的尋找安全、高效、穩(wěn)定的基因載體。相對于病毒載體脂質(zhì)體優(yōu)點(diǎn)在于安全、無免疫源性，缺點(diǎn)是基因轉(zhuǎn)移效率低。因此，脂質(zhì)體的研究主要集中于提高基因轉(zhuǎn)移效率方面。偶聯(lián)了抗體的免疫脂質(zhì)體就是通過抗體的靶向特異性實(shí)現(xiàn)脂質(zhì)體在某些組織器官的聚集來提高基因轉(zhuǎn)移效率。 目的 1．探求將脂質(zhì)體構(gòu)建成為更理想的腦內(nèi)基因轉(zhuǎn)移穿梭載體的方法。2．以神經(jīng)干細(xì)胞表面抗原為靶位，以衍生化單克隆抗體偶聯(lián)脂質(zhì)體腦內(nèi)尋靶，使基因治療更具目的性，提高基因轉(zhuǎn)移效率。 方法 1．使用LB(Luria-Bertani)培養(yǎng)基擴(kuò)增培養(yǎng)pEGFP-C_2質(zhì)粒轉(zhuǎn)化后的JM—109大腸桿菌，以質(zhì)粒抽提試劑盒提取大量pEGFP-C_2質(zhì)粒。經(jīng)過酶切電泳、DNA測序鑒定并以紫外分光光度法檢測質(zhì)粒濃度及純度后用于基因轉(zhuǎn)染。2．以二棕櫚酸磷脂酰乙醇胺(DPPE)對單克隆抗體(anti-Lex／SSEA-1)進(jìn)行衍生化，采用透析法分離純化衍生化抗體、紫外分光光度法檢測抗體衍生化的效率，，間接熒光抗體實(shí)驗(yàn)法檢驗(yàn)衍生化后抗體的活性。實(shí)驗(yàn)中用碳二亞胺法把衍生化的單克隆抗體P-MMA與陽離子脂質(zhì)體DOSPER共價(jià)偶聯(lián)，制成有腦靶向性的
[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

【學(xué)位授予單位】：第一軍醫(yī)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2006
【分類號】：R346

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