两种纯化蓖麻毒素方法的比较

两种纯化蓖麻毒素方法的比较陈兴,赵立超,贺永怀,沈倍奋（军事医学科学院基础医学研究所,北京１００８５０）高纯度的蓖麻毒素（ｒｉｃｉｎ）对于免疫毒素的制备至关重要,直接影响到免疫毒素的应用效果。总结本实验室纯化ｒｉｃｉｎ的经验,我们发现蓖麻粗提物依次通...     

利用重组A蛋白偶联的琼脂糖凝胶纯化α-半乳糖苷酶单克隆抗体

目的:采用一种简单易行的纯化方法获得高纯度α-半乳糖苷酶单克隆抗体。方法:使用重组A蛋白偶联的琼脂糖凝胶捕获和纯化小鼠腹水中的α-半乳糖苷酶单克隆抗体。结果:获得了高纯度、高效价的α-半乳糖苷酶单克隆抗体。结论:应用重组A蛋白偶联的琼脂糖凝胶纯化α-半乳糖苷酶单克隆抗体是一种简单、有效的方法。     

免疫毒素的纯化

本文采用Ultrogel AcA 34凝胶柱纯化单克隆抗体和蓖麻毒素的结合物。结合物与游离毒素的分离较Sephadex G150柱好。继而利用蓖麻毒素B链能与半乳糖基结合的特性,用酸化Sepharose 4B柱进一步纯化结合物;达到完全除去游离抗体,使免疫毒素对靶细胞的特异性毒性提高20倍。     

重组胰高糖素样肽-1融合蛋白的纯化及其活性测定

主要探讨了发酵液中融合蛋白rh（GLP-1A2G）2-HSA的分离纯化过程。发酵上清液经超滤浓缩、离子交换层析、凝胶过滤分离得高纯度的rh（GLP-1A2G）2-HSA。纯化样品经SDS-PAGE检测为单一条带,HPLC分析纯度达98%,^125I标记后用TCA沉淀法测定放射性纯度达97%,符合药效学和药代动力学研究的需要,整个纯化过程回收率达到48.5%。通过细胞增殖实验表明纯化后的rh（GLP-1A2G）2-HSA具有类似GLP-1的促胰岛β细胞增殖活性,说明该分离过程保护了融合蛋白rh（GLP-1A2G）2-HSA的生物活性。通过融合蛋白rh（GLP-1A2G）2-HSA的纯化方法,可获得高纯度的重组融合蛋白rh（GLP-1A2G）2-HSA,为进一步的药效学和药代动力学研究奠定了基础。     

抗人β-BCG单克隆抗体的制备及化学发光检测方法的建立

目的：制备人绒毛膜促性腺激素（β-HCG）单克隆抗体,建立人β-HCG双抗体夹心CLIA检测方法。方法：用人β-HCG抗原免疫小鼠,通过细胞融合、筛选后得到杂交瘤细胞株,然后将细胞株扩大培养并纯化上清液获得抗体,测定抗体亲和力、特异性及表位,最后建立双抗体夹心CLIA方法。结果：获得4株抗人β-HCG的杂交瘤细胞株（β-1—1、β-2—1、β-3—1、β-4—1）。用β-1—1和β-2—1建立的双抗体夹心法的检测范围为0．5mlU／mL．800mlU／mL,灵敏度0．23mIU／mL,检测结果的相对偏差均在±10％内,回收率在90％以上。结论：本研究最终成功制备了抗人β-HCGmAb,建立了定量检测人β-HCG的双抗体夹心CLIA方法,为β-HCG检测及疾病的诊断奠定基础。     

3种纯化重组禽流感病毒NS1抗原方法的比较

目的：摸索出最佳分离纯化和复性重组禽流感病毒NS1抗原的方法,得到高纯度的重组蛋白。方法：将重组质粒pET32a—NS1转染大肠杆菌BL21（DE3）后获得表达,分别以尿素变性、复性,Ni—NTA His．Bind Resin亲和,以及脱氧胆酸钠-N-十二烷基肌氨酸钠（DOC—SKL）洗涤溶解等3种纯化方法从表达产物包涵体中分离纯化NS1蛋白,并进行比较研究。结果：原核表达得到相对分子质量约45000的目的蛋白;3种纯化方法均能分离和纯化出NS1重组蛋白,其中尿素纯化的蛋白纯度为50％～60％,Ni—NTA His．Bind Resin亲和纯化的蛋白纯度为80％-90％,DOC-SKL纯化的蛋白纯度达95％以上;Western blot检测表明,复性后的纯化蛋白具有良好的生物学活性。结论：应用十二烷基肌氨酸钠洗涤纯化是最佳的纯化NS1蛋白的方法,所获得的蛋白可作为包被ELISA的抗原。     

开口箭甾体皂甙元的分离鉴定及其抗荔枝霜疫霉菌活性

在以荔枝霜疫霉菌为指示菌的生物活性测定导向跟踪下,运用TLC、硅胶柱层析、反相硅胶柱层析和凝胶柱层析等分离纯化方法,从开口箭(Tupistra chinensis)甲醇提取物的乙酸乙酯分部萃取物中分离纯化得到2个抗菌化合物。经现代光谱(MS、IR、1D NMR和2D NMR)分析,确定化合物1为1β,2β,3β,4β,5β,7α-六羟基螺甾-25(27)-烯-6-酮,化合物2为螺甾-25(27)-烯-1β,2β,3β,4β,5β,6β,7α-七醇。离体抗菌活性测定化合物1和化合物2抑制荔枝霜疫霉菌(Peronophythora litchii)孢子囊萌发的EC50分别为100.28 mg.mL-1和124.37 mg.mL-1。用质量浓度为500 mg.mL-1的两个化合物分别处理后接种霜疫霉菌,供试荔枝果实的发病率分别为16.7%和18.9%。     

日本血吸虫重组信号蛋白14—3—3的纯化及单克隆抗体的制备

纯化日本血吸虫（中国大陆株）重组信号蛋白（rSj14—3—3）,并制备其单克隆抗体。以纯化后的rsj14-3—3蛋白为抗原免疫Balb／c小鼠,用杂交瘤技术制备抗rSj14-3-3的单克隆抗体,并通过ELISA方法和Westernblotting测定抗体的效价与特异性。获得了大量高纯度的rSj14-3-3蛋白：筛选出了能够稳定分泌抗rSj14．3．3单抗的杂交瘤细胞株3H6。单抗亚型为IgG1。实验依靠大肠杆菌表达系统高效表达了rSj14—3—3蛋白,并利用该蛋白制备了单克隆抗体．可用于今后血吸虫病免疫诊断的实验研究。     

凝胶层析在发酵液中分离提纯透明质酸中的应用

采用葡聚糖凝胶G-100,以0.1 mol/L氯化钠溶液为洗脱剂,在优化的凝胶层析条件(层析柱高度30 cm、进样量2 ml、洗脱流速1 ml/4 min)下,对透明质酸发酵液进行分离纯化,得到了高纯度的透明质酸.HA提取率达到79.85%,蛋白质去除率为84.93%.     

固定pH梯度等电聚焦法分离纯化人胸腺活性肽

 本文用4—7月龄胎儿胸腺,按照匀浆—80℃加热—丙酮沉淀—抽提—超滤等一系列步骤制备人胸腺混合肽。用固定pH梯度聚丙烯酰胺凝胶等电聚焦技术,结合紫外薄层扫描法定位,从胸腺混合肽中得到四种纯化的胸腺肽HTP α_1,β_1,β_2和γ_1;分别测得其等电点为4.6,5.3,5.8和7.6,分子量分别为4800,6700,7200—7300和12,000—13,000。另外至少还得到四种部份纯化的胸腺肽。由玫瑰花结和末端转脱氧核苷酰酶活力测定结果,证明分离得到的各种胸腺肽均有较高的生物活性。     

Curcuminoids as inhibitors of thioredoxin reductase: A receptor based pharmacophore study with distance mapping of the active site

Curcumin is the yellow pigment of turmeric that interacts irreversibly forming an adduct with thioredoxin reductase (TrxR), an enzyme responsible for redox control of cell and defence against oxidative stress. Docking at both the active sites of TrxR was performed to compare the potency of three naturally occurring curcuminoids, namely curcumin, demethoxy curcumin and bis-demethoxy curcumin. Results show that active sites of TrxR occur at the junction of E and F chains. Volume and area of both cavities is predicted. It has been concluded by distance mapping of the most active conformations that Se atom of catalytic residue SeCYS498, is at a distance of 3.56 from C13 of demethoxy curcumin at the E chain active site, whereas C13 carbon atom forms adduct with Se atom of SeCys 498. We report that at least one methoxy group in curcuminoids is necessary for interation with catalytic residues of thioredoxin. Pharmacophore of both active sites of the TrxR receptor for curcumin and demethoxy curcumin molecules has been drawn and proposed for design and synthesis of most probable potent antiproliferative synthetic drugs.     

A Unique Protease Cleavage Site Predicted in the Spike Protein of the Novel Pneumonia Coronavirus (2019-nCoV) Potentially Related to Viral Transmissibility

正Dear Editor,In December 2019, a novel human coronavirus caused an epidemic of severe pneumonia(Coronavirus Disease 2019,COVID-19) in Wuhan, Hubei, China(Wu et al. 2020; Zhu et al. 2020). So far, this virus has spread to all areas of China and even to other countries. The epidemic has caused 67,102 confirmed infections with 1526 fatal cases     

Comparative morphology of the carpel in the Liliaceae: Hewardieae, Petrosavieae, and Tricyrteae

The young pistils in the melanthioid tribes, Hewardieae, Petrosavieae and Tricyrteae, are uniformly tricarpellate and syncarpous. They lack raphide idioblasts. All are multiovulate, with bitegmic ovules. The Petrosavieae are marked by the presence of septal glands and incomplete syncarpy. Tepals and stamens adhere to the ovary in the Hewardieae and the Petrosavieae but not in the Tricyrteae. Two vascular bundles occur in the stamens of the Hewartlieae and Tricyrtis latifolia. Ventral bundles in the upper part of the ovary of the Hewardieae are continuous with compound septal bundles and placental bundles in the lower part. Putative ventral bundles occur in the alternate position in the Tricyrteae and putative placental bundles in the opposite. position in the Petrosavieae. The dichtomously branched stigma in each carpel of the Tricyrteae is supplied by a bifurcated dorsal bundle.     

鸡传染性法氏囊病病毒研究进展

传染性法氏囊病(Infection bursal disease, IBD)是由鸡传染性法氏囊病毒(Infectious bursal disease virus, IBDV)引起的鸡和火鸡的一种高度接触性传染病,给世界各国的禽养殖业带来了巨大损失.自IBDV发现至今新的变异株不断出现,分子结构的改变导致病毒致病力的改变及宿主对疫苗应答的改变,使得传统的疫苗已不能控制其流行,因此各国学者对其基因组结构和功能进行了广泛深入的研究,并积极研制新型有效的疫苗以达到防治的目的.     

Development of a Novel Reverse Transcription Loop-Mediated Isothermal Amplification Method for Rapid Detection of SARS-CoV-2

In conclusion, the novel visual RT-LAMP assay is a simple, rapid, and sensitive approach for detection of SARS-CoV-2, and it is ready for application in primary care and community hospitals or health care centers, and even patients' own houses in response to the current SARS-CoV-2 epidemic because the assay does not require sophisticated equipment and skilled personnel. Furthermore, it is also ready to be used in fields for screening samples from wild animals and environments to facilitate the identification of potential intermediate hosts that mediate the cross-species transmission of SARS-CoV-2 from bats to humans.     

Perinatal Vertical Transmission of Chikungunya Virus in Ruili,a Town on the Border between China and Myanmar

正Dear Editor,Chikungunya virus (CHIKV), an arbovirus in the family of Togaviridae, genus Alphavirus, is transmitted by the A.aegyptii or A. albopictus mosquito, and causes disease in humans characterized by fever, rash, and arthralgia (Silva and Dermody 2017; Suhrbier 2019). It was first reported in 1953 in Tanzania, and caused only a few outbreaks and sporadic cases in Africa and Asia in last century. However, in the epidemic in 2004, CHIKV acquired mutations that conferred enhanced transmission by the A. albopictus mosquito(Schuffenecker et al. 2006). Since then, it has successively caused outbreaks in Africa, the Indian Ocean, South East Asia, the South America, and Europe (Zeller et al. 2016).     

Enterovirus 71 3C proteolytically processes the histone H3 N-terminal tail during infection

Highlights
1. The N-terminal tail of histone H3 is specifically cleaved during EV71 infection.
2. Viral protease 3C is identified as a protease responsible for proteolytically processing the N-terminal H3 tail.
3. Our finding reveals a new epigenetic regulatory mechanism for Enterovirus 71 in virus-host interactions.