糖芯片的制备和检测应用研究进展

近数十年来,糖芯片逐渐成为分析糖介导的识别和结合作用的强有力工具,具有样品检测用量少、特异性强和高通量等优点,可以大大提高糖生物学研究的效率。主要介绍了通过糖库的构建、共价结合和非共价吸附法等方法制备糖芯片的过程,糖芯片的检测方法及其在生物学研究和生物医学领域的应用,以期为糖芯片相关研究提供参考。     

蛋白质芯片构建技术进展

蛋白质芯片是一项实现蛋白质高通量分析和检测的新型技术,在生命科学的各个领域具有重要的应用前景。但是由于蛋白质本身的敏感性质和芯片的独特结构,蛋白质芯片技术的发展面临许多难点,这些正是蛋白质芯片研究者需要共同讨论的问题。本文从芯片构造、探针的固定方法和检测方法三个方面对芯片的构造进行分析和讨论,并对其相关技术的进展进行综述。     

糖芯片研究进展

糖芯片是生物芯片的一种,是继基因芯片、蛋白质芯片、组织芯片等之后发展起来的一种很有前景的生物检测技术。随着糖生物学和糖组学的研究进展,糖芯片正逐步发展为该领域的新型研究手段。介绍了糖芯片技术及其制作方法,高通量分析平台以及糖芯片在生物学研究和医学领域的具体应用,同时也对糖芯片技术的发展进行了展望。     

应用凝集素芯片分析癌细胞膜表面糖链表达

细胞膜表面糖复合物的糖链结构与肿瘤细胞增殖、侵染、转移等发展过程密切相关.凝集素芯片技术的出现实现了对癌症的糖组进行快速、高通量的检测.通过模式细胞系PANC-1证明了构建的凝集素芯片体系的准确性、重复性、特异性,应用这一芯片体系初步检测了几种癌细胞系(HT-29、SGC-7901、BEL-7402、H460)的膜表面糖链表达.这几种癌细胞系表面都有唾液酸、乙酰葡萄糖/葡萄糖、乙酰半乳糖/半乳糖、甘露糖等糖链.根据实验结果,推测它们的细胞膜表面α1-6岩藻糖链表达水平可能较高,而α1-3岩藻糖链表达水平较低;这些聚糖可能是癌症潜在的标志物.凝集素芯片有助于推动癌细胞膜表面糖链的快速分析和筛选出癌症相关的糖链标志物.     

应用凝集素芯片检测肝癌细胞膜表面糖链变化

利用凝集素糖链特异亲和原理构建对细胞膜表面糖链进行即时检测的凝集素芯片体系,检测肝癌发生过程中细胞膜糖链的变化.从H22细胞系、正常小鼠和肝癌模型鼠肝组织中提取细胞进行荧光标记,激光扫描仪检测凝集素位点捕获的细胞,根据凝集素特异亲和性确定细胞膜表面糖表达谱,显微镜下观察捕获细胞的形态.对凝集素芯片捕获细胞的最佳条件进行探讨,用甘露糖抑制试验、流式细胞仪和不同血型红细胞验证了凝集素捕获细胞的特异性.结果显示:正常和肝癌小鼠肝细胞膜表面糖链存在较大差异,正常组只有PSA、DSL、STL、NPL凝集素位点捕获到细胞,实验组只有LTL和DBA位点没有捕获到细胞,提示小鼠肝癌组织细胞膜表面糖链显著增加,细胞膜上唾液酸、乙酰葡萄糖、乙酰半乳糖、甘露糖和半乳糖糖链表达增加,这些糖链及其相关糖蛋白可能在肝癌的发生和发展中起一定作用.该凝集素芯片有较好的稳定性和特异性,可以对细胞膜表面糖链进行动态、即时、通量的检测,为研究细胞膜表面聚糖在细胞发育和癌变等过程中的变化提供了一个技术平台.     

凝集素芯片技术检测糖蛋白方法的建立及初步应用

建立了凝集素芯片技术检测糖蛋白的方法,对实验条件进行了优化,应用凝集素芯片初步检测分析了Chang?蒺s liver正常肝细胞总蛋白中的糖蛋白糖链构成．将凝集素ConA、GNA固定于环氧化修饰的玻片表面,用Cy3标记标准糖蛋白RNaseB,利用凝集素识别特异糖链的原理建立凝集素芯片检测糖蛋白的方法．摸索出最佳封闭剂是含1% BSA的磷酸缓冲液,最佳孵育时间及温度为3 h和室温,最佳孵育缓冲液为含1% BSA和0.05% Tween-20的磷酸缓冲液,并用甘露糖抑制实验验证了凝集素芯片结合的特异性．用包含10种凝集素的芯片,成功解析了标准糖蛋白RNaseB、Fetuin的糖链构成,证实了凝集素芯片检测糖蛋白糖链的可行性．最后用凝集素芯片初步检测分析了Chang?蒺s liver正常肝细胞总蛋白中的糖蛋白糖链构成,发现 Chang's liver正常肝细胞总蛋白中的糖蛋白可能有多价 Sia或GlcNAc、terminalα-1,3 mannose、GalNAc、Galβ-1,4GlcNAc这些糖链结构的存在．蛋白质糖基化是一种重要的翻译后修饰,它在微生物感染、细胞分化、肿瘤转移、细胞癌变等生命活动中起着重要作用,因此近年来蛋白质的糖基化研究受到广泛的重视,但由于缺乏一种简便、快速、高通量的检测手段,蛋白质糖基化修饰的研究发展缓慢．凝集素芯片技术的出现实现了对糖蛋白的快速、准确、高通量的检测 分析．     

凝集素芯片对体液细胞进行检测方法的建立和初步应用

建立对体液细胞进行自动捕获的凝集素芯片体系,利用凝集素对糖链的特异亲和作用捕获细胞,提取白血病患者外周血、肺癌胸水和肝腹水中细胞进行荧光标记,凝集素芯片捕获,激光扫描仪检测捕获细胞的荧光信号,常规HE染色后光学显微镜下观察细胞的形态并进行免疫化学反应,流式细胞仪验证凝集素芯片的特异性．结果表明：凝集素芯片可以对体液中的癌细胞进行自动捕获,对癌细胞膜表面糖链进行识别．芯片检测的细胞浓度最少可达每mL10^4个左右．芯片有较好的重复性和特异性．这种凝集素芯片可用于临床体液中癌细胞的检测分析,对癌细胞膜表面凝集素亲和位点进行即时、高通量的检测,为了解细胞膜表面聚糖在癌变过程中的变化提供了一个技术平台．     

cDNA芯片表面核酸固定化的优化

cDNA芯片技术表面核酸固定化影响因素众多,其中涉及选择载体、固定于玻片的DNA片段浓度、玻片DNA片段的固定方法、玻片预处理方法、DNA片段的变性、溶解DNA片段的点样液等等.针对这些因素进行了优化筛选实验,以便于提高cDNA芯片技术检测基因表达的效率.     

运用树脂加工技术推动生物业务的发展

住友百力的生物开发小组扬眉吐气了。 应用自主树脂加工技术，成功开发出DNA芯片与糖链化工具包。 还将涉足糖链芯片的研究，力争2010年之前使销售额达到100亿日元。[编者按]     

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.     

Recent advances in the study of Kaposi's sarcoma-associated herpesvirus replication and pathogenesis

It has now been over twenty years since a novel herpesviral genome was identified in Kaposi's sarcoma biopsies. Since then, the cumulative research effort by molecular biologists, virologists, clinicians, and epidemiologists alike has led to the extensive characterization of this tumor virus, Kaposi's sarcoma-associated herpesvirus(KSHV; also known as human herpesvirus 8(HHV-8)), and its associated diseases. Here we review the current knowledge of KSHV biology and pathogenesis, with a particular emphasis on new and exciting advances in the field of epigenetics. We also discuss the development and practicality of various cell culture and animal model systems to study KSHV replication and pathogenesis.     

The structure, reproduction and taxonomy of Vidalia and Osmundaria (Rhodophyta, Rhodomelaceae)

Comprises species occurring mostly in subtidal habitats in tropical, subtropical and warm-temperate areas of the world. An analysis of the type species, V. spiralis (Sonder) Lamouroux ex J. Agardh, a species from Australia, establishes basic characters for distinguishing species in the genus. These characters are (1) branching patterns of thalli, (2) flat blades that may be spiralled on their axis, (3) width of the blade, (4) primary or secondary derivation of sterile and fertile branchlets and (5) position of sterile and fertile branchlets on the thalli. Application of the latter two characters provides an important basic method for separation of species into three major groups. Osmundaria , a genus known only in southern Australia, was studied in relation to Vidalia , and its separation from the Vidalia assemblage is not accepted. Species of Vidalia therefore are transferred to the older genus name, Osmundaria. Two new species, Osmundaria papenfussii and Osmundaria oliveae are described from Natal. Confusion in the usage of the epithet, Vidalia fimbriala Brown ex Turner has been clarified, and Vidalia gregaria Falkenberg, described as an epiphyte on Osmundaria pro/ifera Lamouroux, is revealed to be young branches of the host, Osmundaria prolifera.     

New or rare chromosome counts in Artemisia L. (Asteraceae, Anthemideae) and related genera from Kazakhstan

Fifteen chromosome counts of six Artemisia taxa and one species of each of the genera Brachanthemum, Hippolytia, Kaschgaria, Lepidolopsis and Turaniphytum are reported from Kazakhstan. Three of them are new reports, two are not consistent with previous counts and the remainder are confirmations of very scarce (one to four) earlier records. All the populations studied have the same basic chromosome number, x = 9, with ploidy levels ranging from 2x to 6x. Some correlations between ploidy level, morphological characters and distribution are noted.     

肝癌中HBV和HCV基因和抗原的分布及意义

采用原位分子杂交方法检测HCV RNA及HBV X基因;采用免疫组织化学方法研究HCV核心抗原,非结构区C33c抗原及HBxAg在肝细胞肝癌中的定位及分布.结果表明(1)HCV RNA、HBV X基因在肝细胞肝癌组织检出率分别为40%(55/136)和82%(112/136).HCV RNA定位于癌细胞的胞浆内,阳性细胞呈散在、灶状及弥漫分布三种形式;HBV X基因在肝癌细胞中的分布呈胞浆型、核型及核浆型,阳性细胞也呈上述三种分布形式;(2)HCV C33c抗原、核心抗原在肝细胞肝癌中的阳性率为81%(133/164)及86%(141/164).C33c抗原定位于癌细胞及肝细胞的胞浆内;核心抗原既定位于癌细胞核中,又可定位于胞浆中.C33c抗原阳性细胞以灶状分布为主;而核心抗原阳性细     

Selection with two alleles and complete dominance

For a plant selection model with frequency-independent viabilities, fertilities and selfing rates, it is shown that apart from global fixation, for certain parameter combinations a protected polymorphism and facultative fixation (either allele may become fixed according to initial frequencies) may both occur. Facultative fixation requires different selling rates for the dominant and recessive type. Protection of the polymorphism requires resource allocation for male and female function. In this connection the problem of purely genetically caused population extinction is discussed.
For general frequency dependence and regular segregation, the chances for establishment of a completely recessive gene are compared to those of a completely dominant gene. It is proven that the process of establishment of the recessive gene, despite a fitness advantage, may be considerably endangered by drift effects if random mating prevails. The recessive gene may reach the same effectivity in establishment as a dominant gene, only if the recessive homozygote mates exclusively with its own type during the period of establishment.