细胞融合技术

细胞融合是生物工程研究的重要内容和基本技术,综述了细胞融合技术中的常用方法：细胞融合仙台病毒（HVI）诱导法、细胞融合PEG（聚乙二醇）诱导法、细胞融合电场诱导法、细胞融合激光诱导法;以及最新研究进展：基于微流控芯片的细胞融合技术、高通量细胞融合芯片.并对它们的优缺点进行简要的评述.     

细胞融合教学实验的改进

细胞融合技术是研究细胞遗传、基因定位、细胞免疫、病毒和肿瘤的重要手段。所谓细胞融合 ( cell fusion)又称体细胞杂交 ,是指用人工方法使两个或两个以上的体细胞融合成 1个双核或多核细胞 ,不经有性过程而得到杂种细胞。自细胞融合技术 ( Barski,1961)创建至今 ,依据融合过程采用的助融剂的不同可分为 :1)病毒诱导的细胞融合 ;2 )化学因子诱导的细胞融合 ;3 )电场诱导的细胞融合 ,此外还有激光诱导的细胞融合。在保证助融剂及操作技巧的前提下 ,细胞融合成功与否及融合率取决于实验所选择的亲代细胞。目前 ,普通高等院校和高等医学院校…     

电穿透技术在细胞融合和基因转移中的应用

按下电钮,能使一对电极间的细胞融合,还能使基因导入细胞,这是八十年代生物技术中引人注目的新成就之一.电子技术以其精确和简便之特长,又一次推动了生命科学的技术进步.几年前,西德的Zimmermann和Neumann先后报道了分别称作细胞电融合     

草鱼细胞融合及早熟凝集染色体的诱导

用聚乙二醇（PEG）诱导草鱼ZC-7901细胞株融合,测定了不同浓度的PEG诱导草鱼组胞的融合率,从而得出了PEG诱导草鱼细胞融合的适宜浓度为45%（W/W）。用45%PEG诱导间期细胞（I期细胞）与分裂期细胞（M期细胞）融合,早熟凝集染色体（PCC）的诱导率是18.85%。观察PCC形态,G1-PCC呈单股染色体纤维形;G2-PCC呈双股染色纤维形,较分裂期细胞染色体细长;S-PCC呈“粉末状”染色体片段。     

流行性出血热病毒的细胞融合作用

我们的实验结果揭示流行性出血热病毒(EHFV)在酸性条件下可导致感染细胞发生融合,细胞间隙消失,细胞界限不清,细胞和细胞连在一起,形成多核的巨细胞体,姬姆薩染色比未融合细胞浅。融合液(Eagle’s基础培养液,含0.2％BSA,20mmol/L HEPES),用1.0NNaOH调pH至5.0—6.0时,细胞融合最甚,几乎所有的感染细胞都     

病毒介导的细胞融合:癌症发生和发展的新视点

细胞融合(cell fusion)具有重要的生理意义。然而,病毒介导的非生理性细胞融合可能促进癌症的发生和发展。相对于基因突变等细胞癌变的传统诱因,病毒的这种致癌机制能够更合理地解释我们在癌症中发现的许多现象。病毒介导的细胞融合可能诱导癌症的观点对我们重新认识病毒相关肿瘤发生发展的机制,并依此调整癌症治疗的策略具有重要意义。     

以单细胞免疫荧光技术观察烟草细胞融合过程中微管骨架的变化

本文建立了单细胞免疫荧光标记技术并以此结合单对细胞融合技术对细胞融合过程中微管骨架组织形式的动态变化进行了追踪观察。发现在聚乙二醇（PEG）诱导条件下,一旦细胞开始粘连,细胞内微管骨架便开始解聚。在细胞融合的整个过程中一直维持着这种解聚的状态,直到融合完成,在后续的培养中微管骨架才重新出现。在微管骨架呈解聚状态时融合产物不能完成与另外的细胞融合。实验揭示了细胞的再融合能力可能受细胞本身微管骨架状态的影响。该结果为解释高等植物如何避免多精入卵提供了新的可能性。     

固定化细胞技术及其应用研究进展

细胞固定技术是将具有特定生理功能的生物细胞用一定的方法进行固定,并以其作为生物催化剂加以利用的一门技术。相对于游离的单细胞,固定化细胞可简化生产工艺,降低生产成本。本文回顾了细胞固定技术在制备方法和载体材料等方面的研究进展,并总结了近几年来固定化细胞技术在新能源开发、食品加工及环境污染物处理中的应用,对其发展前景进行展望。     

Mating-Type Dependent Cell Pairing in Oxytricha hymenostoma: Macronuclear Fusion and Lack of Meiosis

SYNOPSIS In a hologamy-like process observed in Oxytricha hymenostoma , 2 cells belonging to complementary mating types fused and appeared to contribute equally to the macronuclei of the organism resulting from the fusion. The process was characterized by the absence of micronuclear division and by fusion of the macronuclei of the 2 partners. True conjugation involving micronuclear meiosis was not observed in O. hymenostoma. The biologic significance and the possible adaptive role of the hologamy-like process are discussed.     

Functional Domains within Fusion Proteins: Prospectives for Development of Peptide Inhibitors of Viral Cell Fusion

The entry of enveloped viruses into host cells is accomplished by fusion ofthe viral envelope and target plasma membrane and is mediated by fusionproteins. Recently, several functional domains within fusion proteins fromdifferent viral families were identified. Some are directly involved inconformational changes after receptor binding, as suggested by the recentrelease of crystallographically determined structures of a highly stablecore structure of the fusion proteins in the absence of membranes. However,in the presence of membranes, this core binds strongly to the membrane'ssurface and dissociates therein. Other regions, besides the N-terminal fusionpeptide, which include the core region and an internal fusion peptide inparamyxoviruses, are directly involved in the actual membrane fusion event,suggesting an umbrella like model for the membrane inducedconformational change of fusion proteins. Peptides resembling these regionshave been shown to have specific antiviral activity, presumably because theyinterfere with the corresponding domains within the viruses. Overall, thesestudies shed light into the molecular mechanism of membrane fusion induced byenvelope glycoproteins and suggest that fusion proteins from different viralfamilies share common structural and functional motifs.     

体外受精研究：玉米和烟草性细胞融合的几种模式

分别用玉米 (ZeamaysL .)和烟草 (NicotianatabacumL .)为材料 ,以单对细胞融合技术广泛开展了各种组合的性细胞的融合实验。重点观察了性细胞融合过程中细胞体积对细胞膜融合行为的影响。视频增差显微术结合细胞膜荧光标记技术显示 ,性细胞融合过程中受融合双方体积差异大小的调节 ,细胞膜的融合行为可归纳为 3种基本模式 ;发现在体外受精条件下精细胞可以类似被吞噬的方式进入卵细胞 ,且其融合动作非常迅速。由此提出一种假设机制用以解释体内受精过程中雄性细胞质被排除的可能原因。     

体外受精研究:玉米和烟草性细胞融合的几种模式

分别用玉米(Zea mays L.)和烟草(Nicotiana tabacum L.)为材料,以单对细胞融合技术广泛开展了各种组合的性细胞的融合实验.重点观察了性细胞融合过程中细胞体积对细胞膜融合行为的影响.视频增差显微术结合细胞膜荧光标记技术显示,性细胞融合过程中受融合双方体积差异大小的调节,细胞膜的融合行为可归纳为3种基本模式;发现在体外受精条件下精细胞可以类似被吞噬的方式进入卵细胞,且其融合动作非常迅速.由此提出一种假设机制用以解释体内受精过程中雄性细胞质被排除的可能原因.     

Fusion protein technologies for biopharmaceuticals: Applications and challenges

Stefan R. Schmidt consolidates the hugely diverse field of fusion proteins and their application in the creation of biopharmaceuticals. The text is replete with case studies and clinical data that inform and intrigue the reader as to the myriad possibilities available when considering the creation of a fusion protein. This valuable text will serve the novice as a broad introduction or the seasoned professional as a thorough review of the state of the art. The first marketed therapeutic recombinant protein was human insulin (Humulin® R). Its approval in 1982 was followed by other such products, including erythropoietin (EPO), interferon (IFN), and tissue plasminogen activator (tPa). Since the 1980s, the number and general availability of recombinant products that replace natural proteins harvested from animal or human sources has increased considerably. Following the initial success, researchers started de novo designs of therapeutic proteins that do not occur in nature. The first of these new drugs to be approved was etanercept (Enbrel®), a fusion portion containing a section of the tumor necrosis factor (TNF) receptor fused to the Fc portion of human IgG1.     

proUK-KGDW融合基因在CHO细胞中的高表达

利用常规分子生物学技术,构建了新型高效的proUK-KGDW融合基因的分泌型哺乳动物细胞表达载体。将该载体线性化后转染CHO/dhfr-细胞,经G418筛选获得阳性克隆,然后挑取表达水平较高的克隆进行MTX加压扩增,以提高proUK-KGDW杂合体的表达水平,经2～3轮MTX加压扩增,获得多株表达水平超过10μg/(106细胞·24h)的稳定的高表达细胞株,为proUK-KGDW杂合体的制备及功能研究奠定了基础。     

Fusion of cultured dog kidney (MDCK) cells: I. Technique,fate of plasma membranes and of cell nuclei

Summary The evaluation of the intracellular signal train and its regulatory function in controlling transepithelial transport with electrophysiological methods often requires intracellular measurements with microelectrodes. However, multiple impalements in epithelial cells are hampered by the small size of the cells. In an attempt to avoid these problems we fused cells of an established cell line, Madin Darby canine kidney cells, originally derived from dog kidney, to giant cells by applying a modified polyethylene glycol method. During trypsin-induced detachment from the ground of the petri dish, individual cells grown in a monolayer incorporate volume and mainly lose basolateral plasma membrane by extrusion. By isovolumetric cell-to-cell fusion, spherical giant cells are formed within 2 hr. During this process a major part of the individual cell plasma membranes is internalized. Over three weeks following cell plasma membrane fusion degradation of single cell nuclei and cell nuclear fusion occurs. We conclude that this experimental approach opens the possibility to investigate ion transport of epithelia in culture by somatic cell genetic techniques.     

Fusion protein technologies for biopharmaceuticals: Applications and challenges: Editor Stefan R Schmidt

Stefan R. Schmidt consolidates the hugely diverse field of fusion proteins and their application in the creation of biopharmaceuticals. The text is replete with case studies and clinical data that inform and intrigue the reader as to the myriad possibilities available when considering the creation of a fusion protein. This valuable text will serve the novice as a broad introduction or the seasoned professional as a thorough review of the state of the art. The first marketed therapeutic recombinant protein was human insulin (Humulin® R). Its approval in 1982 was followed by other such products, including erythropoietin (EPO), interferon (IFN), and tissue plasminogen activator (tPa). Since the 1980s, the number and general availability of recombinant products that replace natural proteins harvested from animal or human sources has increased considerably. Following the initial success, researchers started de novo designs of therapeutic proteins that do not occur in nature. The first of these new drugs to be approved was etanercept (Enbrel®), a fusion portion containing a section of the tumor necrosis factor (TNF) receptor fused to the Fc portion of human IgG1.