本文選題：免疫磁性微球 + 殼聚糖　； 參考：《暨南大學(xué)》2005年碩士論文

【摘要】：免疫磁性微球(Immunomagnetic microspheres簡(jiǎn)稱IMM)是將單抗或者生物活性物質(zhì)通過(guò)共價(jià)交聯(lián)或物理吸附包被到有磁性高分子復(fù)合微球的表面,形成具有免疫活性的磁性微球。免疫磁性分離是基于細(xì)胞表面抗原以及某種蛋白與連有磁性微球的特異性單抗相結(jié)合,在外加磁場(chǎng)的條件下,通過(guò)抗體與IMM相連的細(xì)胞和目標(biāo)蛋白被吸附而滯留在磁場(chǎng)中,由于微球表面的單抗只結(jié)合含有特異性抗原的細(xì)胞和蛋白,從而達(dá)到與其他種類的細(xì)胞和蛋白相分離的目的。免疫磁性分離的方法與常用的細(xì)胞分離方法(FACS法、二抗致敏紅細(xì)胞法、與組織培養(yǎng)管吸附的抗體結(jié)合法等)相比,有方便、快速、低毒、高效的優(yōu)點(diǎn),而迅速滲透到免疫、生理、病理、藥理、微生物等領(lǐng)域。本文從殼聚糖磁性復(fù)合微球的制備出發(fā),在以下幾個(gè)方面取得一定的進(jìn)展: 1.用反相微乳的方法合成了CMCM(Chitosan magnetic composite microspheres簡(jiǎn)稱CMCM),對(duì)粒徑和表觀形貌、微球鐵含量、磁響應(yīng)性、表面官能基團(tuán)等性質(zhì)進(jìn)行了表征,與單體聚合法制備的聚苯乙烯磁性復(fù)合微球(Polystyrene magnetic composite microspheres,簡(jiǎn)稱PMCM)進(jìn)行比較后得知,CMCM是比PMCM更理想的免疫磁性微球載體材料,有可能成為一種蛋白吸附性能良好的磁性高分子復(fù)合微球。 2.探討了不同條件下CMCM的蛋白吸附條件,在此基礎(chǔ)上初步探討了免疫磁性微球的制備和細(xì)胞分離富集的可行性�；罨蟮腃MCM對(duì)小牛血清蛋白(BSA)的吸附性能較活化前提高了近5倍(從5.4mg/g到24.6mg/g)。時(shí)間對(duì)吸附量影響甚微,溫度對(duì)吸附行為影響也很小,離子濃度對(duì)活化的磁性微球的吸附有一定的影響,隨著NaCl濃度逐漸增加至1.0mol/l,微球?qū)SA的吸附逐漸降低,因此應(yīng)盡量避免在離子濃度較高的條件下進(jìn)行吸附操作。 3.把制備的CMCM經(jīng)過(guò)戊二醛的活化,再連接鼠抗人CD_3單抗制備了CD_3免疫磁性微球,用所制備鼠抗人CD_3單抗的免疫磁性微球可以特異性地從人全血中分離出CD_3~+細(xì)胞,分離純度經(jīng)流式細(xì)胞儀檢測(cè)均達(dá)到95%以上。證明了CMCM作為免疫磁性微球載體的可行性。
[Abstract]:Immunomagnetic microspheres (imm) is a kind of immunoreactive magnetic microspheres, which is coated on the surface of magnetic polymer composite microspheres by covalent crosslinking or physical adsorption of monoclonal or bioactive substances. Immunomagnetic separation is based on the binding of a cell surface antigen and a protein to a specific monoclonal antibody attached to a magnetic microsphere. Under the condition of an external magnetic field, cells and target proteins linked to imm by antibodies are adsorbed in the magnetic field. Because the McAbs on the surface of the microspheres only bind to the cells and proteins containing specific antigens, they can be separated from other kinds of cells and proteins. The method of immunomagnetic separation is more convenient, faster, less toxic and more efficient than the usual methods of cell separation, second antibody sensitized red blood cell, antibody binding to tissue culture tube, and so on. Physiology, pathology, pharmacology, microbiology and other fields. Based on the preparation of chitosan magnetic composite microspheres, some progress has been made in the following aspects: 1. CMCM-chitosan magnetic composite microspheres was synthesized by inverse microemulsion method. The particle size and morphology, iron content, magnetic response and surface functional groups were characterized. Compared with the polystyrene magnetic composite microspheres, prepared by monomer polymerization, the results show that the polystyrene magnetic composite microspheres are more ideal than PMCMs in immunomagnetic microspheres. It is possible to become a kind of magnetic polymer composite microspheres with good protein adsorption performance. 2. The protein adsorption conditions of CMCM under different conditions were discussed, and the feasibility of preparation and cell separation and enrichment of immunomagnetic microspheres was preliminarily discussed. The adsorption performance of activated CMCM for bovine serum protein (BSA) was nearly five times higher than that before activation (from 5.4mg/g to 24.6 mg / g). Time had little effect on the adsorption amount and temperature had little effect on the adsorption behavior. Ion concentration had a certain effect on the adsorption of activated magnetic microspheres. With the increase of NaCl concentration to 1.0 mol / l, the adsorption of BSA on the microspheres gradually decreased. Therefore, the adsorption operation should be avoided under the condition of high ion concentration as far as possible. 3. The prepared CMCM was activated by glutaraldehyde and then connected with mouse anti-human CDS3 monoclonal antibody to prepare CDS3 immunomagnetic microspheres. CD3 cells could be specifically isolated from human blood by using immunomagnetic microspheres prepared by mouse anti human CD3 McAb. The purity was more than 95% by flow cytometry. The feasibility of CMCM as a carrier of immunomagnetic microspheres was proved.
【學(xué)位授予單位】：暨南大學(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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