人凝血酶原复合物离子交换树脂的筛选及吸附性能研究

采用若干种人工合成的大孔阴离子交换树脂,对人血浆中凝血酶原复合物(PCC)的分离纯化性能进行了研究.同时.将DEAE-Sephadex A50时PCC的分离纯化性能与该类人工合成分离纯化材料进行了对比.研究结果表明,在各种人工合成大孔阴离子交换树脂中,CG-6对PCC具有较优的分离纯化性能;并系统研究了CG-6树脂对PCC分离纯化性能.     

Beta proteobacterium sp.T1菌株产壳聚糖酶的分离纯化

目的:分离纯化Beta proteobacterium sp.T1菌株所产的壳聚糖酶.方法:采用(NH4)2SO4(20%～70%)分级盐析、阴离子交换树脂DEAE Cellulose 52柱层析、SephadexG-100柱层析技术进行分离纯化,采用SDS-PAGE鉴定酶的纯度、分子量.结果:经DEAE Cellulose 52柱层析,壳聚糖酶纯化了13.19倍;经Sephadex G-100柱层析,壳聚糖酶纯化了26.32倍.结论:纯化后的酶经SDS-PAGE鉴定已达到电泳纯,分子量29.5kDa.     

应用单克隆抗体免疫亲和柱纯化干细胞因子

应用抗人干细胞生长因子(SCF)单克隆抗体,通过化学偶联方法制备成亲和层析柱,用于纯化大肠杆菌表达的可溶性重组人rh-SCF.结果表明,经亲和柱纯化的样品经SDS-PAGE电泳检测纯度达95%以上,计算蛋白回收率为23.4%,并且纯化后rh-SCF生物活性明显高于纯化前样品.     

凝胶过滤与镍柱亲和纯化金葡菌α-溶血素重组蛋白的生物学特性比较

以在宿主菌株BL21(DE3)中成功表达的重组金黄色葡萄球菌α-溶血素蛋白为研究对象,分析比较通过凝胶过滤层析(Gel filtration chromatography)和镍柱亲和层析纯化试剂盒(Ni-NTA spin columns)纯化所得到的重组蛋白的蛋白含量和生物特性方面的差异.SDS-PAGE分析检测纯化产物,Bradford法测定蛋白含量,兔红细胞测定半数溶血效价,结果显示这2种方法得到的纯化产物在53 kD处均呈现单一清晰带,达电泳级纯度.与此同时,凝胶过滤对目的蛋白的纯化率为14.04%,蛋白含量为0.337 mg/mL,溶血活性为1519 HU/mg;镍柱亲和层析的纯化率为17.5%,蛋白含量为0.35 mg/mL.溶血活性为1463 HU/mg.由此可见,凝胶过滤得到的纯化产物在蛋白含量和蛋白活性方面丝毫不亚于镍柱亲和层析纯化试剂盒.     

豌豆铁蛋白的纯化及其抗血清的制备

干豌豆种子粗提物经MgCl2 盐析、AcA2 2 凝胶过滤和DEAE 纤维素阴离子交换柱层析等方法进行纯化 ,以邻菲咯啉显色法检测铁蛋白 ,最后获得纯的铁蛋白 .纯化的铁蛋白在PAGE上显示一条带 ,SDS PAGE显示该蛋白仅含 2 8kD一条亚基 .纯化的豌豆铁蛋白免疫兔 7周后 ,琼脂糖双扩散法检测抗血清效价达 1∶32 .用分级盐析法纯化抗血清 ,纯化后的抗体用琼脂糖双扩散法对大豆铁蛋白粗提物有免疫交叉反应     

抗体分离纯化技术的研究进展

制备高特异性、高效价的抗体是实验免疫学技术的基础,抗体质量的高低,将直接影响试验的成败.抗体的制备有两个途径:一是一般通用的方法,以纯化的抗原免疫动物,获得多克隆抗体;二是应用杂交瘤技术制备单克隆抗体.但不论用何种技术制备的抗体都需要进行纯化.重要的是根据抗体的性质和来源选择一个合适的分离纯化方法.对当前的纯化方法进行了一个简要的综述.     

重组蛋白质亲和标签的选择与串联亲和纯化的应用进展

重组蛋白质技术作为蛋白质研究的重要手段之一,在生物化学与生物物理学研究领域,扮演着极其重要的角色.亲和纯化作为最为方便与快捷的重组蛋白质纯化手段,日益得到广泛的应用.由于各种亲和标签,纯化介质层出不穷,性质各异.应根据自身研究对象具体情况选择合适的亲和纯化标签.近年双亲和标签进行串联亲和纯化日益成为蛋白质相互作用研究的重要方法.多种亲和标签的搭配已得到成功应用,部分已进入商业化,并在各种模式生物中得到广泛的应用,本文就重组蛋白质亲和标签的选择与串联亲和纯化作一综述.     

PSCA-HSP70融合蛋白在大肠杆菌中的表达与纯化

旨在大肠杆菌中表达PSCA-HSPT0融合蛋白,并对其进行纯化.克隆人PSCA基因及HSP70基因,构建表达PSCA-HSP融合蛋白的工程菌,优化表达及纯化条件,对重组蛋白进行纯化.结果表明,成功构建重组表达质粒PSCA-HSP,重组蛋白得到可溶性表达,优化纯化条件后获得90%以上纯度的重组蛋白.本研究成功实现了PSCA与HSP的融合表达,为下一步肿瘤疫苗的研制奠定基础.     

亲和层析法分离鉴定人类端粒酶复合体及其蛋白质组分分析

为分离纯化人类端粒酶复合体并对其蛋白质组分进行分析 ,自行设计一套特异的以端粒重复序列为配体 ,以亲和磁珠为介质 ,以竞争性碱基序列为洗脱原则的寡核苷酸亲和纯化法 ,对He La细胞端粒酶复合体进行分离纯化 .并采用近年发展起来的主要用于检测序列特异性 DNA结合蛋白的凝胶迁移阻滞分析法对纯化产物进行鉴定分析 .应用 SDS- PAGE对纯化蛋白质亚基组分进行分析与鉴定 ,并采用电泳迁移率和已知蛋白质分子质量标准的对数作图分析测得所得蛋白质亚基成分的相对分子质量 .结果表明 ,纯化产物以 TRAP法检测酶活性可见典型的梯形条带 ;比活性为每 mg蛋白质的 cpm值为 1 80× 1 0 9,纯化倍数 1 80 0 ,得率 90 % .凝胶迁移阻滞法分析显示特异的凝胶迁移阻滞性电泳条带 ;以 SDS- PAGE检测得到 4种蛋白质亚基成分 ,其蛋白质相对分子质量分别为 2 2 0 ku、2 1 2 ku、1 1 6ku和 43ku.由上述可见 ,采用自行设计的寡核苷酸亲和纯化法获得了人端粒酶复合体及 4种蛋白质亚基组分 .     

抗人DR5多克隆抗体的制备与纯化

目的:分离纯化融合蛋白GST-eDR5,免疫小鼠制备抗人DR5多克隆抗体.方法:用GST纯化试剂盒和电泳两次纯化的融合蛋白GST-eDR5免疫小鼠,制备抗CST-eDR5抗血清,然后用GST纯化得到抗人DR5抗血清.通过Westem blot、ELISA方法鉴定抗血清特异性和效价.结果:通过亲和纯化得到了高纯度的融合蛋白GST-eDR5,其浓度为0.65μg/μl.免疫产生的DR5抗血清特异性高,且效价高达1:25600,纯化后的抗人DR5抗血清不再识别GST,只识别人DR5.结论:成功制备了高特异性、高效价的抗人DR5多克隆抗体,为深入研究其对肿瘤细胞的生长抑制和凋亡作用提供了实验工具.     

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.