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小鼠抗寨卡病毒包膜蛋白胞外区(Eecto)单克隆抗体的制备

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细胞与分子免疫学杂志 2024年第5期40卷 447-454页

作者：薛潘董阳超武福星陈洋张剑元航武苏闪袁若栋李宝莉雷迎峰延安大学医学院陕西延安716000 空军军医大学基础医学院微生物与病原生物学教研室陕西西安710032

目的制备小鼠抗寨卡病毒(ZIKV)包膜蛋白胞外区(Eecto)单克隆抗体。方法首先构建ZIKV Eecto的原核表达质粒pET28a-ZIKV-Eecto,将其转化至大肠杆菌感受态BL21后,利用异丙基β-D-硫代半乳糖苷(IPTG)诱导表达。重组Eecto蛋白以包涵体形式表... 详细信息

目的制备小鼠抗寨卡病毒(ZIKV)包膜蛋白胞外区(Eecto)单克隆抗体。方法首先构建ZIKV Eecto的原核表达质粒pET28a-ZIKV-Eecto,将其转化至大肠杆菌感受态BL21后,利用异丙基β-D-硫代半乳糖苷(IPTG)诱导表达。重组Eecto蛋白以包涵体形式表达,经过变性、复性和超滤获得纯化的蛋白。将Eecto蛋白3次免疫后,获得多克隆抗体血清,进行效价测定,并利用ZIKV prME真核表达质粒转染的HEK293T细胞进行Western blot法和免疫荧光法分析。取效价较高的小鼠脾脏制备脾细胞,与SP2/0骨髓瘤细胞融合,通过有限稀释法获取分泌抗体的杂交瘤细胞,并进一步制备腹水。对腹水进行抗体效价测定、亚类分析。以ZIKV同属病毒日本脑炎病毒(JEV)、黄热病病毒(YFV)、登革病毒1-4(DENV1-4)、蜱传脑炎病毒(TBEV)的Eecto为包被抗原,利用ELISA分析ZIKV单克隆抗体的交叉反应性,进一步对效价高、特异性强的抗体进行特异性分析。结果成功构建了原核表达质粒pET28a-ZIKV-Eecto,并获得纯化的Eecto蛋白,该蛋白具有良好的免疫原性;制备筛选得到1D6、4F11、4H7、4F8四株单克隆抗体,其中三株(1D6、4H7、4F8)为IgGκ型抗体,一株(4F11)为IgMκ,1D6腹水效价大于1∶108;其中1D6和4H7为ZIKV特异性抗体,与其他黄病毒无交叉反应。结论成功制备了小鼠抗ZIKV Eecto单克隆抗体,为后续建立ZIKV检测方法及致病机制研究提供了实验材料。

关键词：寨卡病毒(ZIKV) 包膜蛋白胞外区(Eecto) 原核表达单克隆抗体

小鼠抗西方马脑炎病毒E2蛋白胞外区(E2ecto)单克隆抗体的制备

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细胞与分子免疫学杂志 2024年第1期40卷 62-68页

作者：武福星董阳超张剑薛潘袁若栋陈洋元航李宝莉雷迎峰延安大学医学院陕西延安716000 空军军医大学基础医学院微生物与病原生物学教研室陕西西安710032

目的制备针对西方马脑炎病毒(WEEV)E2包膜糖蛋白胞外区(E2ecto)的小鼠特异性单克隆抗体(mAb)。方法构建表达WEEV E2ecto的原核表达质粒pET-28a-WEEV E2ecto,转化BL21(DE3)感受态细胞后,IPTG诱导表达,主要为包涵体形式。包涵体纯化后,通... 详细信息

目的制备针对西方马脑炎病毒(WEEV)E2包膜糖蛋白胞外区(E2ecto)的小鼠特异性单克隆抗体(mAb)。方法构建表达WEEV E2ecto的原核表达质粒pET-28a-WEEV E2ecto,转化BL21(DE3)感受态细胞后,IPTG诱导表达,主要为包涵体形式。包涵体纯化后,通过变性、复性、超滤等制备得到E2ecto蛋白;以QuickAntibody-Mouse5W为佐剂,将E2ecto蛋白肌肉免疫BALB/c小鼠,间隔21 d加强1次。免疫后35 d,取效价超过1×104的小鼠腹腔接种1次E2ecto蛋白,3 d后取小鼠脾细胞与SP2/0细胞融合。通过有限稀释法筛选稳定分泌特异性mAb的杂交瘤细胞,接种小鼠腹腔后制备腹水;利用ELISA检测抗体亚型及腹水中抗体效价的水平;将真核表达质粒pCAGGS-WEEV-CE3E2E1转染BHK-21细胞,通过免疫荧光法(IFA)鉴定上述mAb的生物学活性;利用东方马脑炎病毒(EEEV)、委内瑞拉马脑炎病毒(VEEV)的E2ecto蛋白进行ELISA鉴定上述抗体的特异性。结果成功表达及复性获得了纯化的WEEV E2ecto蛋白;制备了3G6G10、3D7G2、3B9E8、3D5B7四株mAb,其亚型分别为IgG2c(κ)、IgM(κ)、IgM(κ)、IgG1(κ);3G6G10、3B9E8、3D7G2腹水抗体效价均为105,3D5B7达到107;该4株抗体与VEEV和EEEV等其他脑炎甲病毒无交叉反应。结论成功制备了4株特异性结合WEEVE2ecto的小鼠mAb。

关键词：西方马脑炎病毒(WEEV) E2包膜糖蛋白胞外区(E2ecto) 原核表达单克隆抗体(mAb)

An overview of cetacean strandings, bycatches and rescues along the western coast of the Taiwan Strait, China: 2010–2015

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Acta Oceanologica Sinica 2017年第12期36卷 31-36页

作者： ZHAO Liyuan ZHU Qian MIAO Xing XU Min wu fuxing DAI Yufei TAO Cuihua MOU Jianfeng WANG Xianyan Laboratory of Marine Biology and Ecology Third Institute of OceanographyState Oceanic Administration Ocean College Shandong University (Weihai)

The analysis of cetacean strandings can provide fundamental information about species diversity and composition in a particular region. The present study collected and analyzed cetacean strandings, bycatches and rescues along the western coast of the Taiwan Strait, China, from 2010 to 2015. In total, 48 records, including 37 strandings, 8 bycatches and 3 rescues, involving 13 known species were collected. Among them, Indo-Pacific humpback dolphins（Sousa chinensis） and finless porpoises（Neophocaena phocaenoides） were the most common species, consisting of 31.3% and 25%, respectively. Notably, 10 out of the 48（20.8%） records were collected from the Pingtan Island and included 3 species of Mysticeti and 4 species of Odontoceti. Finally, we compared the cetacean species composition between the western and eastern coasts of the Taiwan Strait; 31 cetacean species occurred in the Taiwan Strait, indicating a relatively high cetacean diversity in this region. Systematic field surveys are urgently needed to explore the cetacean species composition, population stock and the related habitat status in the Taiwan Strait, which may improve conservation management in the future.

关键词： cetacean stranding bycatch rescue species diversity Taiwan Strait

Early divergence and differential population histories of the Indo-Pacific humpback dolphin in the Pacific and Indian Oceans

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Integrative Zoology 2021年第4期16卷 612-625页

作者： Liyuan ZHAO Watchara SAKORNWIMON Wenzhi LIN Peijun ZHANG Rachawadee CHANTRA Yufei DAI Reyilamu AIERKEN fuxing wu Songhai LI Kongkiat KITTIWATTANAWONG Xianyan WANG Laboratory of Marine Biology and Ecology Third Institute of OceanographyMinistry of Natural ResourcesXiamenChina Marine and Coastal Resources Research Center The Middle Gulf of ThailandChumphonThailand Institute of Deep-sea Science and Engineering Chinese Academy of SciencesSanyaChina Marine and Coastal Resources Research Center The Upper Gulf of ThailandSamut SakhonThailand Fujian Provincial Key Laboratory of Marine Ecological Conservation and Restoration XiamenChina Phuket Marine Biological Research Center PhuketThailand

The currently recognized Indo-Pacific humpback dolphin occurs in estuaries and surrounding shallow waters from the South China Sea to the Asian coast of the Indian ***,a recent study suggested that the humpback dolphin from the Bay of Bengal may represent a distinct phylogenetic *** this study,we sequenced 915-bp mtDNA segments from five geographic populations in both Chinese and Thai waters;together with previously published sequences,these data revealed that the ancestral Indo-Pacific humpback dolphin might have split during the transition from the Oligocene to Miocene(23.45 Mya,95%HPD:16.65–26.55 Mya),and then dispersed along the Pacific and Indian Ocean coasts of *** differentiation was detected between most of the examined populations,except for only a few pairwise populations in the northern South China *** differentiation/distance between the humpback dolphins from the northern and southern South China Sea met the sub-species threshold value proposed for marine mammals,whereas that between the humpback dolphins in the Pacific and the Indian Ocean was above the species *** inference of historic gene flow indicated low but constant northward gene flow along the Indian Ocean coast;however,there was a recent abrupt increase in gene flow in the Pacific region,likely due to the shortening coastline at the low stand of sea *** results revealed that the current taxonomic classification of Indo-Pacific humpback dolphins may not reflect their phylogeography.

关键词： historic gene flow Indo-Pacific region phylogenetic species concept phylogeography Sousa chinensis

Impacts of global climate change on birds and marine mammals in Antarctica

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Advances in Polar Science 2017年第1期28卷 1-12页

作者： wu fuxing DONE Lu ZHANG Yanyun ZHANG Zhengwang Ministry of Education Key Laboratory for Biodiversity Sciences and Ecological Engineering College of Life SciencesBeijing Normal University Beijing 100875 China

Birds and marine mammals in Antarctica, especially penguins and seals, are top consumers and critical elements of the Southem Ocean ecosystem. As a region undergoing rapid global change, new challenges will be posed to the survival of these vertebrates species. Global climate change causes many alterations, such as ocean temperature rise, altered sea ice distribution, and abnormal climate events along with effects of intensive human activities, such as fishing. These not only directly affect the spatiotemporal distributions and population dynamics of Antarctic birds and marine mammals but also indirectly influence them via modification of their food resources. At present, the impact of climate change on birds and marine mammals in the Antarctica is focusing on a number of species in a few areas. Response mechanisms of these species are still very limited and therefore require further long-term and continuous monitoring and research.

关键词： Antarctica birds marine mammal global climate change

Seasonal group characteristics and occurrence patterns of Indo-Pacific humpback dolphins(Sousa chinensis) in Xiamen Bay,Fujian Province,China

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Seasonal group characteristics and occurrence patterns of In...

RPA-毛细管电泳检测人腺病毒55型的方法建立及初步应用

福建省海洋学会2016年学术年会

作者： Xianyan Wang fuxing wu Samuel T.Turvey Massimiliano Rosso Qian Zhu Laboratory of Marine Biology and Ecology Third Institute of OceanographyState Oceanic Administration

Monthly field surveys were conducted between August 2010 and July 2015 to explore seasonal group characteristics and occurrence patterns of Indo-Pacific humpback dolphins(Sousa chinensis) in Xiamen Bay,*** formed larger groups in winter and spring(dry seasons)than in summer and autumn(wet seasons)(U = 1564.00,P wuyu,and Dadeng-Xiaodeng) during the wet *** occurrence patterns may be associated with the seasonal prey shifts between these different environments.

关键词： Sousa chinensis seasonal variation group characteristics occurrence pattern Xiamen Bay

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空军军医大学学报 2024年第1期45卷 66-72页

作者：袁若栋董阳超武福星段甜薛潘张剑袁明城薛智丰张海军张欠欠雷迎峰高小朋延安大学生命科学学院陕西延安716000 空军军医大学基础医学院病原生物学与微生物学教研室陕西西安710032

目的建立重组酶聚合酶扩增(RPA)技术联合毛细管电泳快速检测人腺病毒55型(HAdV55)的方法及初步应用。方法构建HAdV55-Hexon质粒作为扩增模板,针对HAdV55保守区Hexon基因序列设计RPA特异性引物,对引物筛选和优化,并建立RPA扩增及联合毛... 详细信息

目的建立重组酶聚合酶扩增(RPA)技术联合毛细管电泳快速检测人腺病毒55型(HAdV55)的方法及初步应用。方法构建HAdV55-Hexon质粒作为扩增模板,针对HAdV55保守区Hexon基因序列设计RPA特异性引物,对引物筛选和优化,并建立RPA扩增及联合毛细管电泳检测体系,进行了特异性、灵敏度、可靠性评价;对18份临床样本进行RPA毛细管电泳检测。结果构建了含有HAdV55-Hexon全长基因的质粒,获得一对扩增效率最高的RPA引物;HAdV55的RPA毛细管电泳检测体系最低检出限为2.9×10^(3) copies/mL,整个过程30 min即可完成,冠状病毒(HCoV-229E、HCoV-OC43、HCoV-HKU1)、甲型乙型流感病毒、副流感病毒、呼吸道合胞病毒标准品提取物用该体系检测时均为阴性,且检测常见的人腺病毒类型(HAdV3、HAdV4、HAdV7、HAdV16、HAdV21)时也均为阴性,可认为对HAdV55有较强的特异性检测;建立的RPA毛细管电泳检测能够有效地检测出临床患者咽拭子中的HAdV55。结论RPA联合毛细管电泳检测HAdV55方法的建立为其临床诊断提供了良好的基础。

关键词：人腺病毒55型重组酶聚合酶扩增毛细管电泳

Social organization and distribution patterns inform conservation management of a threatened Indo-Pacific humpback dolphin population

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Social organization and distribution patterns inform conserv...

Evocation (Gan-Xing) as a Core Proposition in the Aesthetics of the Wei and Jin Dynasties: An Analysis of the Rhapsodies on Thoughts (Gan-Fu)

福建省海洋学会2016年学术年会

作者： Xianyan Wang fuxing wu Samuel T.Turvey Massimiliano Rosso Cuihua Tao Xiaohui Ding Qian Zhu Third Institute of Oceanography State Oceanic Administration

From August 2010 to December 2013,field photo-identification surveys were conducted to explore the social organization and distribution patterns of a threatened Indo-Pacific humpback dolphin(Sousa chinensis) population in Xiamen Bay,*** of 58 photo-identified dolphins,56 dolphins that were sighted at least 2 times were used for social structure *** association pattern among individuals was divided into 2 discrete communities with strong geographic adherence:27 dolphins assigned to an eastern community(EC) and 29 dolphins assigned to a western community(WC).Among the WC individuals,9 dolphins formed mixed groups with EC individuals at Tongan Bay and the mouth of Tongan Bay,leading to low-level intercommunity *** 50%kernel density estimate(KDE) and minimum convex polygon(MCP) of each community were calculated,with the core habitats presented here differing substantially from those reported in previous *** propose that the geographical segregation of these 2 communities and the apparent shift in core habitats could be the result of adaptation by dolphins to local environmental conditions,notably intensive anthropogenic activities.

关键词： community segregation distribution pattern Indo-Pacific humpback dolphin social organization Xiamen Bay

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Frontiers of Literary Studies in China-Selected Publications from Chinese Universities 2024年第3期18卷 314-335页

作者： XUE fuxing College of Philosophy Nankai UniversityTianjin 300350China

The rhapsodies on the mind (xin-fu) is a major category of the rhapsody (fu) genre during the Wei and Jin dynasties. Those describing human sentimental psychology are known as the rhapsodies on thoughts (gan-fu). The rhapsodies on thoughts from this period embody a special proposition on how human aesthetic experience arises, which is termed as evocation (gan-xing). Evocation involves two dimensions: responding to objects (gan-wu) and responding to seasons (gan-shi). Together, they form the basic framework or mechanism that generates the abstract aesthetic experience in the rhapsodies on thoughts. More specifically, the rhapsodies on thoughts reveal five types of aesthetic experience generated: (1) aesthetic emotions triggered by natural sights;(2) philosophical insights triggered by natural sights;(3) aesthetic emotions triggered by social environments;(4) aesthetic experience triggered by natural sights and social environments;and (5) anti-evocation experience, namely an aesthetic response to the natural world triggered by specific scenes in life. These five evocation models broadly cover the aesthetic experience generated by the rhapsodies on thoughts from this period. First and foremost, evocation reveals the internal mechanism that generates aesthetic experience—external things evoke one’s inner feelings, thus creating aesthetic appreciation. Furthermore, it uncovers the normative significance of natural aesthetic appreciation as the prerequisite for artistic aesthetic appreciation: People are first moved by external objects in the realm of natural aesthetic appreciation before consciously developing the urge to express their emotions or feelings through natural images in the realm of artistic aesthetics. The rise of evocation, originating from the rhapsodies on thoughts of the Wei and Jin dynasties, signifies that Chinese aesthetic consciousness was attained during this period in relation to the mechanism of generating aesthetic experiences.

关键词： evocation responding to objects responding to seasons aesthetic consciousness