ERα(sc-542)抗体说明书 santa
SANTA CRUZ BIOTECHNOLOGY,INC.ER α(MC-20):sc-542Santa Cruz Biotechnology,Inc. 1.800.457.3801831.457.3800fax 831.457.3801Europe +BACKGROUNDEstrogen receptors (ER)are members of the steroid/thyroid hormone receptor superfamily of ligand-activated transcription factors.Estrogen receptors,including ER αand ER β,contain DNA binding and ligand binding domains and are critically involved in regulating the normal function of reproductive tissues.ER αand ER βhave been shown to be differentially activated by vari-ous ligands.Receptor-ligand interactions trigger a cascade of events,including dissociation from heat shock proteins,receptor dimerization,phosphorylation and the association of the hormone activated receptor with specific regulatory elements in target genes.Evidence suggests that ER αand ER βmay be regulated by distinct mechanisms even though they share many functional characteristics.REFERENCES1.Danielian,P .S.,et al.1992.Identification of a conserved region required for hormone dependent transcriptional activation by steroid hormone receptors.EMBO J.11:1025-1033.2.Mosselman,S.,et al.1996.ERb:identification and characterization of a novel human estrogen receptor.FEBS Lett.392:49-53.CHROMOSOMAL LOCATIONGenetic locus:ESR1(human)mapping to 6q25.1;ESR1(mouse)mapping to 10A1.SOURCEER α(MC-20)is an affinity purified rabbit polyclonal antibody raised against a peptide mapping at the C-terminus of ER αof mouse origin.PRODUCTEach vial contains 200µg IgG in 1.0ml of PBS with <0.1%sodium azide and 0.1%gelatin.Blocking peptide available for competition studies,sc-542P ,(100µg peptide in 0.5ml PBS containing <0.1%sodium azide and 0.2%BSA).Available as TransCruz reagent for Gel Supershift and ChIP applications,sc-542X,200µg/0.1ml.Available as fluorescein (sc-542FITC)or rhodamine (sc-542TRITC)conjugates for immunofluorescence,200µg/ml.Available as Alexa Fluor ®405(sc-542AF405),Alexa Fluor ®488(sc-542AF488)or Alexa Fluor ®647(sc-542AF647)conjugates for immunofluores-cence;100µg/2ml.Alexa Fluor ®is a trademark of Molecular Probes,Inc.,Oregon,USASTORAGEStore at 4°C,**DO NOT FREEZE**.Stable for one year from the date of shipment.Non-hazardous.No MSDS required.RESEARCH USEFor research use only,not for use in diagnostic procedures.APPLICATIONSER α(MC-20)is recommended for detection of estrogen receptor αof mouse,rat and human origin by Western Blotting (starting dilution 1:200,dilution range 1:100-1:1000),immunoprecipitation [1-2µg per 100-500µg of total protein (1ml of cell lysate)],immunofluorescence and immunohistochemistry (including paraffin-embedded sections)(starting dilution 1:50,dilution range 1:50-1:500)and solid phase ELISA (starting dilution 1:30,dilution range 1:30-1:3000).Suitable for use as control antibody for ER αsiRNA (h):sc-29305,ER αsiRNA (m):sc-29306,ER αsiRNA (h2):sc-44204,ER αsiRNA (r):sc-45949ER αshRNA Plasmid (h):sc-29305-SH,ER αshRNA Plasmid (m):sc-29306-SH,ER αshRNA Plasmid (h2):sc-44204-SH,ER αshRNA Plasmid (r):sc-45949-SHER αshRNA (h)Lentiviral Particles:sc-29305-V,ER αshRNA (m)Lentiviral Particles:sc-29306-V,ER αshRNA (h2)Lentiviral Particles:sc-44204-V and ER αshRNA (r)Lentiviral Particles:sc-45949-V.ER α(MC-20)X TransCruz antibody is recommended for Gel Supershift and ChIP applications.Molecular Weight of ER α:66kDa.DATASELECT PRODUCT CITATIONS1.Massaad-Massade,L.,et al.2002.HMGA1enhances the transcriptional activity and binding of the estrogen receptor to its responsive element.Biochemistry 41:2760-2768.2.Guo,Z.,et al.2002.Estradiol-induced nongenomic calcium signaling regu-lates genotropic signaling in macrophages.J.Biol.Chem.277:7044-7050.3.McCarthy,T.L.,et al.2003.Runx2integrates estrogen activity in osteo-blasts.J.Biol.Chem.278:43121-43129.4.Perissi,V.,et al.2004.A corepressor/coactivator exchange complex required for transcriptional activation by nuclear receptors and other regulated transcription factors.Cell 116:511-526.5.Omoto,Y.,et al.2005.Estrogen receptor αand imprinting of the neonatal mouse ventral prostate by A 102:1484-1489.6.Pedram,A.,et al.2007.A conserved mechanism for steroid receptor translocation to the plasma membrane.J.Biol.Chem.282:22278-22288.7.Klein,C.,et al.2010.Transcriptional profiling of equine endometrium during the time of maternal recognition of pregnancy.Biol.Reprod.83:102-113.ER α(MC-20):sc-542.Western blot analysis of human recombinant ER α.<ERα116K -80K -52K -35K -30K-ER α(MC-20):sc-542.Immunoperoxidase staining of formalin-fixed,paraffin-embedded human breast carcinoma tissue showing nuclear localization.。
A群链球菌抗原检测试剂盒使用说明书
A群链球菌抗原检测试剂盒（胶体金法）使用说明书【产品名称】通用名称：A群链球菌抗原检测试剂盒（胶体金法）英文名称：Strep A Test【包装规格】25人份/盒。
【预期用途】A群链球菌抗原检测试剂盒（胶体金法）是一种快速免疫层析试验，用于定性检测咽拭子标本中的A群化脓性链球菌抗原。
A群链球菌是引起咽炎的主要病原体。
对该致病原做出准确诊断是正确治疗这种疾病的重要条件。
链球菌感染后要选用抗生素治疗，如果未经治疗，可能会引起诸如风湿热之类的严重后遗症1。
检测A群链球菌感染的传统方法是将咽拭子标本培养24-48小时后，证实β溶血菌落是A群链球菌2。
阳性结果可通过观察羊血琼脂平板上微生物对杆菌肽的敏感性而获得3，4。
基于碳水化合物抗原群特异性可对A群链球菌做出免疫学诊断5。
A群链球菌抗原检测试剂盒（胶体金法）使用的是免疫层析方法，所用抗体对A群链球菌是特异的。
试验设计可对抗原在试剂上进行提取，没有转移步骤，标本完全包含在检测卡中，结果6分钟可得。
【检验原理】A群链球菌抗原检测试剂盒（胶体金法）是一种检测咽拭子标本中A群化脓性链球菌抗原的薄膜免疫层析试验。
一条吸附到硝酸纤维素膜上的兔抗A群链球菌抗体带，作为样本线，另一条吸附到同一膜上的兔抗种属抗体,作为对照线。
结合有可视粒子的兔抗A群链球菌抗体和种属抗体干燥结合到惰性纤维支持物上，形成的结合物垫与带条带的薄膜结合在一起构成检测条。
检测条和一个放咽拭子的小孔位于书形检测卡的两侧。
试验时，将咽拭子插入检测卡中，从滴瓶中加入提取试剂，将咽拭子转三圈。
温育1分钟后，将检测卡闭合，使提取的标本与检测条接触。
A群链球菌抗原被固定的A群链球菌抗体捕获，并与结合物抗体结合。
固定的抗种属抗体捕获另一可视结合物，阳性结果5分钟内出现，5分钟读数时的阴性结果表明不存在A群链球菌抗原。
试验结果可用紫红色线的存在与否来解释。
阳性结果会出现样本线和对照线两条线，阴性结果只出现一条对照线，其他检测结果表示试验无效。
注射用阿替普酶使用说明书(德国进口)
注射用阿替普酶使用说明书(德国进口)之答禄夫天创作中文名：注射用阿替普酶商品名：爱通立曾用名：艾通立英文名：Actilyse化学名称：Plasminogen activator (human tissue-type protein moiety)分子式:CHNOPS分子量:C2569H3894N746O781S40活性成分:阿替普酶 Alteplase （重组人组织纤维卵白溶酶原激活剂 (rt-PA)）性状:本品为白色至类白色冻干粉末, 无嗅.除活性成分外, 还含有以下辅料：精氨酸、磷酸、吐温80及注射用水.20毫克包装盒内有一个含20毫克活性成分（干粉总重933毫克）的小瓶及一个内装20毫升注射用水的注射用小瓶.50毫克包装盒内有一个含50毫克活性成分（干粉总重2333毫克）的小瓶及一个内装50毫升注射用水的注射用小瓶.作用机理:本药的活性成分是一种糖卵白, 可直接激活纤溶酶原转化为纤溶酶.当静脉给予时, 本品在循环系统中暗示出相对非活跃状态, 一旦与其纤维卵白结合后, 本品被激活, 诱导纤溶酶原转化为纤溶酶, 招致纤维卵白降解, 血块溶解.心肌梗死的研究：在一项入选了40000多例急性心肌梗死患者的研究（GUSTO）中, 治疗组给予100毫克本品90分钟滴注, 静脉滴注肝素辅助治疗, 30天死亡率为6.3% ；对比组予以150万单元链激酶60分钟滴注, 辅以皮下或静滴肝素比较, 其30天死亡率为7.3%.同时本品治疗的患者溶栓后60分钟和90分钟的血管再通率高于链激酶治疗的患者.在180分钟及以后时间的血管再通率2组没有不同.本品治疗的患者比未经溶栓治疗的患者30天死亡率低.与未经溶栓疗法的患者相比, 本品治疗的患者总体心室功能及局部心肌壁运动功能受损较轻.一项抚慰剂对比试验（LATE）标明, 发病后6-12小时内给予治疗, 经本品100 mg 3小时治疗的患者30天死亡率比对比组低.对某些呈现明显心肌梗死症状的病例, 发病后24小时内的溶栓治疗也有益处.肺栓塞的研究：急性年夜面积肺栓塞伴血流动力学不稳定的患者使用本品溶栓, 可迅速缩小血栓, 并降低肺动脉压.无死亡率的资料.急性缺血性脑卒中的研究：在2项美国试验（NINDS A/B）中, 与抚慰剂相比, 使用本品预后良好（无功能缺陷或轻微功能缺陷）的患者比例明显增高.在2个欧洲试验和另外1个美国试验中未能证实上述发现, 然而在这后3个试验中, 年夜部份患者未能在脑卒中发作的3小时内接受治疗.随后的分析标明在脑卒中发作3小时内接受本品治疗的疗效是肯定的.尽管严重的和致命性的颅内出血的风险增高, 但与抚慰剂相比, 预后良好的差值为14.9%（95%可信区间为8.1-21.7%）.此数据无法给出关于治疗对死亡率影响简直切结论.然而总体来说, 如果在卒中症状发作的3小时内给予本品且遵循本说明书描述的各注意事项, 应用本品的收益还是年夜于可能的风险的.随后进行的对所有临床数据的分析显示, 与症状发作3小时内就给予本品治疗的患者相比, 在症状发作3小时后（3-6小时）才接受本品治疗的患者疗效差, 而且风险增高, 这招致其收益/风险比值落在原来0-3小时内可接受的值以外.由于本品具有纤维卵白相对特异性, 100毫克的本品可能招致循环中纤维卵白原在4小时内减少至60%左右, 但一般24小时后可恢复到80%以上.4小时后纤溶酶原和α-2-抗纤溶酶分别减少至20%和35%左右, 24小时后可恢复到80%以上.只有少数患者呈现明显的较长时间的循环纤维卵白原水平下降.毒理研究:在年夜鼠和南美猴的亚急性毒理研究中, 未发现其它预期之外的不良反应.致突变试验中未发现有致突变倾向.药代动力学:本品可从血循环中迅速清除, 主要经肝脏代谢（血浆清除率550-680毫升/分钟）.相对血浆α半衰期（T1/2α）是4～5分钟, 这意味着20分钟后, 血浆中本品的含量不到最初值的10%.周边室的残留量, 其β半衰期为40分钟.适应症：急性心梗、血流不稳定的急性年夜面积肺栓塞、急性缺血性脑卒中的溶栓治疗.急性心肌梗死：对症状发生6小时以内的患者, 采用90分钟加速给药法（拜会用法用量）；对症状发生6-12小时以内的患者, 采用3小时给药法（拜会用法用量）.本品已被证实可降低急性心肌梗死患者30天死亡率.血流不稳定的急性年夜面积肺栓塞：可能的情况下应借助客观手段明确诊断, 如肺血管造影或非侵入性手段如肺扫描等.尚无证据显示对与肺栓塞相关的死亡率和晚期发病率有积极作用.急性缺血性脑卒中：必需预先经过恰当的影像学检查排除颅内出血之后, 在急性缺血性脑卒中症状发作后的3小时内进行治疗.用法用量：心梗发病6 hr内：采用90分钟加速给药法.静推15 mg, 其后30分钟内静滴50 mg, 剩余的35 mg在60分钟内静滴, 直至最年夜剂量达100 mg.心梗发病6-12 hr ：采用3小时给药法.静推10 mg, 其后1 hr内静滴50 mg, 剩余的按10 mg/30分钟, 至3小时末静滴完毕, 直至最年夜剂量达100 mg.体重< 65 kg患者, 给药总剂量不应超越1.5 mg/kg.肺栓塞：应在2 hr内给予100 mg, 10 mg 在1-2分钟内静推, 90 mg在2 hr内静滴.急性缺血性脑卒中：在卒中症状发作后的3 hr内尽快给予0.9 mg/kg, 最年夜剂量为90 mg.先将剂量的10%静脉推入, 剩余剂量在1 hr内静滴.（详见说明书）应在症状发生后尽快按以下指导剂量给药.无菌条件下將一小瓶爱通立干粉（10、20或50毫克）用注射用水溶解为1毫克/毫升或2毫克/毫升的浓度.使用爱通立20毫克或50毫克包装中的移液套管完成上述溶解把持.如果是爱通立10毫克, 则使用注射器.10 毫克规格：加入干粉中注射用水10毫升, 终浓度为1毫克/毫升；加入干粉中注射用水5毫升, 终浓度为2毫克/毫升.20 毫克规格：加入干粉中注射用水20毫升, 终浓度为1毫克/毫升；加入干粉中注射用水10毫升, 终浓度为2毫克/毫升.50 毫克规格：加入干粉中注射用水50毫升, 终浓度为1毫克/毫升；加入干粉中注射用水25毫升, 终浓度为2毫克/毫升.配制好的溶液应通过静脉给药.配制的溶液可用灭菌生理盐水(0.9%)进一步稀释至0.2毫克/毫升的最小浓度.心肌梗死:对在症状发生6小时以内的患者, 采用90分钟加速给药法：15 mg静脉推注, 终浓度为1毫克/毫升的溶液15毫升, 或终浓度为2毫克/毫升的溶液7.5毫升.随后30分钟继续静脉滴注50 mg, 终浓度为1毫克/毫升的溶液50毫升, 或终浓度为2毫克/毫升的溶液25毫升.剩余35 mg于60分钟继续静脉滴注, 终浓度为1毫克/毫升的溶液35毫升, 或终浓度为2毫克/毫升的溶液17.5毫升.直至最年夜剂量达100 mg.体重在65公斤以下的患者, 给药总剂量应按体重调整, 具体如下：15 mg静脉推注, 终浓度为1毫克/毫升的溶液15毫升, 或终浓度为2毫克/毫升的溶液7.5毫升.然后按0.75毫克/公斤体重的剂量继续静脉滴注30分钟（最年夜剂量50毫克）, 终浓度为1毫克/毫升的溶液0.75毫克/公斤体重, 或终浓度为2毫克/毫升的溶液0.375毫克/公斤体重.剩余的按0.5毫克公斤/体重的剂量继续静脉滴注60分钟（最年夜剂量35毫克）, 终浓度为1毫克/毫升的溶液0.5毫克/公斤体重, 或终浓度为2毫克/毫升的溶液0.25毫克/公斤体重.对在症状发生6-12小时以内的患者, 采用3小时给药法：10 mg静脉推注, 终浓度为1毫克/毫升的溶液10毫升, 或终浓度为2毫克/毫升的溶液5毫升.其后1小时继续静脉滴注50 mg, 终浓度为1毫克/毫升的溶液50毫升, 或终浓度为2毫克/毫升的溶液25毫升.剩余剂量每30分钟静脉滴注10毫克, 终浓度为1毫克/毫升的溶液10毫升/30分钟, 或终浓度为2毫克/毫升的溶液5毫升/30分钟.至3小时末滴完, 最年夜剂量为100 mg.体重在65公斤以下患者, 给药总剂量不应超越 1.5 mg/kg体重.本品最年夜剂量为100毫克.辅助治疗：症状发生后尽快给予阿司匹林, 并在心肌梗死发生后的第1个月内继续给药.推荐剂量为160 - 300 mg/天.同时给予肝素24小时或更长时间（加速给药法中至少应陪伴给药48小时）.建议在溶栓治疗前静脉注射5000国际单元, 然后以1000国际单元/小时继续静脉滴注.肝素剂量应根据反复测定的aPTT值调整, aPTT值应为用药前的1.5 - 2.5倍.肺栓塞:本品100 mg应继续2小时静脉滴注.最经常使用的给药方法为：10 mg在1-2分钟内静脉推注, 终浓度为1毫克/毫升的溶液10毫升, 或终浓度为2毫克/毫升的溶液5毫升.90 mg在随后2小时继续静脉滴注, 终浓度为1毫克/毫升的溶液90毫升, 或终浓度为2毫克/毫升的溶液45毫升.体重缺乏65公斤的患者, 给药总剂量不应超越 1.5 mg/kg体重.辅助治疗：静滴本品后, 当aPTT值低于正常上限2倍时, 应给予（或再次给予）肝素.静滴肝素剂量应根据aPTT值调整, aPTT值应为用药前的1.5-2.5倍.急性缺血性脑卒中:治疗必需由神经科医师进行（拜会禁忌和注意事项）.推荐剂量为0.9 mg/kg体重（最年夜剂量为90 mg）.总剂量的10%先从静脉推入, 剩余剂量在随后60分钟继续静脉滴注.治疗应在症状发作后的3小时内开始.辅助治疗：在症状发生的最初24小时内, 此治疗方案与肝素和阿司匹林合用的平安性和有效性尚未进行系统研究.在本品治疗后的24小时以内应防止使用阿司匹林或静脉给予肝素.若给予肝素以防治其它症状（如防止深静脉栓塞发生）, 则剂量不得超越10000国际单元, 并由皮下注射给药.药物过量:尽管本品具有相对纤维卵白特异性, 但过量后仍会呈现显著的纤维卵白原及其它凝血因子的减少.年夜大都情况下, 停用本品治疗后, 生理性再生足以弥补这些因子.然而, 如发生严重的出血, 建议输入新鲜冻干血浆或新鲜全血, 如有需要可使用合成的抗纤维卵白溶解剂.禁忌:本品不成用于有高危出血倾向者, 如已知出血体质；口服抗凝药, 如华法令；目前或近期有严重的或危险的出血；已知有颅内出血史或疑有颅内出血；疑有蛛网膜下腔出血或处于因动脉瘤而招致蛛网膜下腔出血状态；有中枢神经系统病变史或创伤史（如肿瘤、动脉瘤以及颅内或椎管内手术）；最近（10天内）曾进行有创的心外按压、分娩或非压力性血管穿刺（如锁骨下或颈静脉穿刺）；严重的未获得控制的动脉高血压；细菌性心内膜炎或心包炎；急性胰腺炎；最近3个月有胃肠溃疡史、食管静脉曲张、动脉瘤或动脉/静脉畸形史；出血倾向的肿瘤；严重的肝病, 包括肝功能衰竭、肝硬变、门静脉高压（食管静脉曲张）及活动性肝炎；最近3个月内有严重的创伤或年夜手术.治疗急性心肌梗死时的弥补禁忌：有脑卒中史.治疗急性肺栓塞时的弥补禁忌：有脑卒中史.治疗急性缺血性脑卒中时的弥补禁忌：缺血性脑卒中症状发作已超越3小时尚未开始静脉滴注治疗或无法确知症状发作时间；开始静脉滴注治疗前神经学指征缺乏或症状迅速改善；经临床（NIHSS>25）和/或影像学检查评定为严重脑卒中；脑卒中发作时陪伴癫痫发作；CT扫描显示有颅内出血迹象；尽管CT扫描未显示异常, 仍怀疑蛛网膜下腔出血；48小时内曾使用肝素且凝血酶原时间高于实验室正常值上限；有脑卒中史并陪伴糖尿病；近3个月内有脑卒中发作；血小板计数<100 × 109/L ；收缩压>185毫米汞柱或舒张压>110毫米汞柱, 或需要强力（静脉内用药）治疗手段以控制血压；血糖<2.8 mmol/L（50 mg/dl）或>22.2 mmol/L（400 mg/dl）.儿童及老年患者用药：本品不能用于18岁以下及80岁以上的急性脑卒中患者治疗注意事项:必需有足够的监测手段才华进行溶栓/纤维卵白溶解治疗.只有经过适当培训且有溶栓治疗经验的医生才华使用本品, 而且需有适当的设备来监测使用情况.建议在备有标准复苏装置和药物的地址使用爱通立进行治疗.老年患者颅内出血的危险增加, 因此, 对老年患者应仔细权衡使用本品的风险及收益.到目前为此, 本品用于儿童的经验还很有限.如同其他所有溶栓剂, 应该慎重权衡预期治疗收益和可能呈现的危险, 特别是对以下患者：较小的近期损伤, 如活组织检查、主要血管的穿刺、肌肉注射及心外按压；在禁忌中未曾提及的出血倾向.防止使用硬质导管.治疗急性心肌梗死时的弥补注意事项：本品的用量不应超越100毫克否则颅内出血的发生率可能增高.应确保本品的剂量遵从本说明书中用法用量的规定.本品重复用药的经验有限, 使用本品一般不引起过敏反应.如发生过敏样反应, 应停止滴注本品并给予相应的治疗.应该慎重权衡预期治疗收益和可能呈现的危险, 特别是对收缩压高于160毫米汞柱的患者.治疗急性肺栓塞时的弥补注意事项：同急性心肌梗死治疗.治疗缺血性脑卒中时的弥补注意事项：特别注意, 只有神经专科已经过培训的且有经验的医师才华进行相应治疗.特殊注意事项, 收益/风险比可能下降：与治疗其他适应症相比, 本品用于急性缺血性脑卒中治疗时颅内出血的风险明显增加, 因为出血主要发生在梗塞部位.需特别注意以下情况：所有禁忌中包括的事项以及所有可能增加出血风险的情况；微小的尚无症状的脑动脉瘤；预先经阿司匹林治疗且症状发生后没有及时给予本品治疗的患者可能有更年夜的脑出血的风险.在这种情况下, 本品的用量不得超越0.9毫克/公斤体重（最年夜剂量90毫克）.如果症状发生已超越3小时, 则患者不得再用本品治疗（拜会禁忌）, 因为不良的收益/风险比值主要基于以下情况：随着时间推移, 预期的阳性治疗效果会下降；预先经阿司匹林治疗的患者其死亡率增加；症状性出血的风险增加.临床经验证明, 应当在治疗过程中进行血压监测且需延长至24小时.如果收缩压超越180毫米汞柱或舒张压高于105毫米汞柱, 建议进行静脉内抗高血压治疗.在有卒中史或其糖尿病未获得控制的患者, 本品治疗的获益降低, 可是这些患者仍然可以从治疗中受益.对非常轻度的脑卒中患者, 治疗风险超越收益（拜会禁忌）.对非常严重的脑卒中患者, 其脑出血风险和死亡率均增高（拜会禁忌）, 不得使用本品治疗.广泛性梗塞的患者其预后不良的风险很高, 包括可能呈现严重出血和死亡.对这些患者, 应仔细权衡收益/风险比.随着患者年龄、脑卒中严重性和血糖水平的增高, 其预后良好的可能性下降而发生严重功能缺陷、死亡或脑出血的可能性增加, 与治疗自己无关.年龄80岁以上, 严重脑卒中（经临床诊断或影像学诊断）及血糖基础值小于50 mg/dl或年夜于400 mg/dl的患者不得使用本品治疗（拜会禁忌）.其它特殊注意事项：缺血部位的再灌注可诱发梗塞区域的脑水肿.由于可能招致出血风险增加, 在本品溶栓后的24小时内不得使用血小板聚集抑制剂治疗.儿童用药:目前, 儿童使用本品的经验还很有限.老年患者用药:拜会注意事项.孕妇及哺乳期妇女用药:妊娠和哺乳妇女使用本品的经验非常有限.对急性的危及生命的疾病, 应权衡收益与潜在危险.静脉给予药理上的有效剂量对妊娠植物无致畸作用.每天按超越3毫克/公斤体重给药, 可诱发家兔胚胎毒性反应（胚胎死亡、发育迟滞）.剂量超越每日10毫克/公斤体重, 对年夜鼠围产期发育或生殖参数没有影响.目前尚不知晓爱通立是否能够泌入乳汁.不良反应:概述：使用本品后的最罕见不良反应是出血, 招致红细胞比积和/或血红卵白下降.与溶栓治疗相关的出血可分成2种类型：概况出血, 常为穿刺部位或血管损伤处出血；内出血, 为胃肠道、泌尿生殖道、后腹膜、中枢神经系统或实质脏器出血.死亡和永久残疾的陈说见于发生卒中（包括颅内出血）和其他严重出血事件的患者.以下不良反应（根据MedDRA系统脏器分类列表）是在与爱通立治疗可能有关的不良事件陈说基础上获得.以下所列不良事件的发生频率得自于一项包括8299例患者的使用爱通立治疗心肌梗死的临床研究.胆固醇结晶栓塞形成的分类未见于临床试验人群, 而基于自发性陈说数据所得.临床试验中, 因肺栓塞和脑卒中（治疗时间窗：发作后0-3小时）而入选接受治疗的患者数量远少于上述因心肌梗死而入选的患者数量.因此, 与来自年夜样本的心肌梗死研究的数据相比, 那些数据缺乏明显不同可能是因样本较小所致.除治疗脑卒中时发生的颅内出血和治疗心肌梗死时发生的再灌注后心律失常, 没有医学依据能够假设爱通立在治疗肺栓塞和急性缺血性脑卒中时的副作用与其治疗心肌梗死时所发生的副作用在数量和水平上有所分歧.很罕见：>10% ；罕见：>1%但≤10% ；不罕见：>0.1%但≤1% ；罕见：>0.01%但≤0.1% ；非常罕见：≤0.01%.心肌梗死适应症：心血管系统异常 - 很罕见再灌注后心律失常, 可能危及生命并需要惯例抗心律失常治疗.心肌梗死和肺栓塞适应症：神经系统异常 - 不罕见颅内出血.急性缺血性脑卒中适应症：神经系统异常 - 罕见颅内出血, 以症状性脑内出血为主（可多至10%的患者）.但并未显示整体致残率或死亡率因此上升.心肌梗死、肺栓塞和急性缺血性脑卒中适应症：免疫系统异常 - 不罕见过敏样反应, 通常为轻度, 但在个别病例可危及生命.其暗示可以是皮疹、荨麻疹、支气管痉挛、血管源性水肿、低血压、休克和其他与过敏反应有关的症状.一旦呈现上述异常, 应给予惯例抗过敏治疗.在发生过敏样反应的患者中, 有相对较年夜比例者同时服用血管紧张素转换酶抑制剂.目前尚不知晓确定的针对爱通立的过敏反应（IgE介导）.在极少数病例中曾观察到一过性、低滴度爱通立抗体形成, 但无法确立与之相关的临床意义.血管异常- 很罕见出血；罕见瘀斑；不罕见血栓栓塞, 可招致相关脏器发生相应后果；罕见实质脏器出血, 胆固醇结晶栓塞, 可招致相关器官发生相应后果；非常罕见眼出血.呼吸、胸部与纵隔异常 - 罕见鼻出血.胃肠道异常 - 罕见出血至胃肠道管腔内、恶心、呕吐.恶心和呕吐也是心肌梗死发作时的症状；不罕见腹膜后出血, 牙龈出血.肾脏与尿道异常 - 罕见泌尿生殖道出血.全身和注射部位异常 - 很罕见体表出血, 通常发生于穿刺处或血管损伤部位.未分类的不良反应- 很罕见血压下降；罕见体温升高.需要手术和其他医学处置 - 罕见需要输血.关于某些严重或罕见事件的其他信息：在临床试验中, 当根据临床惯例治疗患者, 亦即不成急诊左心导管拔出术, 则患者偶需输血.如果有潜在的出血危险尤其是脑出血, 则应停止溶栓治疗.因本品的半衰期短, 对凝血系统影响轻微, 所以一般不用给予凝血因子.年夜大都出血患者, 可经中断溶栓和抗凝治疗, 扩容及人工压迫损伤血管来控制出血.如在出血发生的4小时内已使用肝素, 则应考虑使用鱼精卵白.对少数使用守旧治疗无效的患者, 可输注血制品, 包括冷沉淀物, 新鲜冻干血浆和血小板, 每次使用后应做临床及实验室的再次评估.纤维卵白原水平为1克/升时可输注冷沉淀物.抗纤维卵白溶解剂可作为最后一种治疗选择.心肌梗死或肺栓塞患者可能会经历与疾病相关的其它不良事件如心力衰竭、再缺血、心绞痛、心跳停止、心源性休克、再梗塞、瓣膜功能异常（如主动脉瓣破裂）和肺栓塞.在溶栓治疗中这些不良事件也曾有报道, 可能有生命危险甚至招致死亡.具有同类药理学活性物质的不良反应：与其它溶栓剂相同, 个别病例报道发生中枢神经系统事件（如惊厥）, 这些事件通常与发生的缺血性或出血性脑血管事件有关.药物相互作用:在应用本品治疗前、治疗同时或治疗后24小时内使用香豆素类衍生物、口服抗凝剂、血小板聚集抑制剂、普通肝素、低分子肝素和其它影响凝血的药物可增加出血危险（拜会禁忌）.同时使用血管紧张素转换酶抑制剂可能增加过敏样反应的危险.在呈现如此反应的患者中, 有年夜部份患者正在同时使用血管紧张素转换酶抑制剂的治疗.配伍禁忌：配制的溶液可用灭菌生理盐水（0.9%）按1:5稀释.可是不能继续使用注射用水或用碳水化合物注射液如葡萄糖对配制的溶液作进一步稀释.本品不能与其它药物混合, 既不能用于同一输液瓶也不能应用同一输液管道（肝素亦不成以）.FDA妊娠分级:C级: 植物研究证明药物对胎儿有危害性（致畸或胚胎死亡等）, 或尚无设对比的妊娠妇女研究, 或尚未对妊娠妇女及植物进行研究.本类药物只有在权衡对孕妇的益处年夜于对胎儿的危害之后, 方可使用.贮藏/有效期:保管于原始包装中.避光, 低于25°C贮存.溶液配制后, 推荐立即使用.已经证实配制好的溶液能够在2-8°C坚持稳定24小时, 勿冷冻.有效期:36个月MIMS药物分类:抗凝血、抗血小板和纤维卵白溶解剂(Anticoagulants, Antiplatelets & Fibrinolytics (Thrombolytics))制造商:勃林格殷格翰 (Boehringer Ingelheim)代办署理/经销商:国药集团药业股份有限公司 (CNCM Pharm)。
酿酒酵母 Hansenula polymorpha 中表达乙肝病毒 PreS2-S 抗原说明书
© WIV, CAS and Springer-Verlag Berlin Heidelberg 2014D ECEMBER 2014 V OLUME 29 I SSUE 6 403R ESEARCH ARTICLEGeneration of Hepatitis B virus PreS2-S antigen in Hansenula polymorphaXiaowei Xu 1,2, Sulin Ren 2, Xiaoxiao Chen 2, Jun Ge 2, Zhenxing Xu 2, Hongying Huang 2, Honglin Sun 2, Yue Gu 2, Tong Zhou 2, Jianqiang Li 2✉, Hanmei Xu 1✉1. State Key Laboratory of Nature Medicines, China Pharmaceutical University, Nanjing 210009, China2. Institute of Vaccine Research, Jiangsu Simcere Pharmaceutical Group, Nanjing 210042, ChinaDespite the long availability of a traditional prophylactic vaccine containing the HBV surface antigen (HBsAg) and aluminum adjuvant, nearly 10% of the population remains unable to generate an effective immune response. Previous studies have indicated that hepatitis B virus (HBV) PreS2-S is abundant in T/B cell epitopes, which induces a stronger immune response than HBsAg, particularly in terms of cytotoxic T lymphocyte (CTL) reaction. In the current study, the HBV PreS2-S gene encoding an extra 26 amino acids (PreS2 C-terminus) located at the N-terminus of HBsAg was cloned into the pVCH1300 expression vector. PreS2-S expressed in the methylotrophic yeast, Hansenula polymorpha , was produced at a yield of up to 250 mg/L. Subsequent puri fi cation steps involved hydrophobic adsorption to colloidal silica, ion-exchange chromatography and density ultracentrifugation. The fi nal product was obtained with a total yield of ~15% and purity of ~99%. In keeping with previous studies, ~22 nm virus-like particles were detected using electron microscopy. The generated PreS2-S antigen will be further studied for ef fi cacy and safty in animals. KEYWORDS hepatitis B virus (HBV); PreS2-S; virus-like particle (VLP); Hansenula polymorphaReceived: 27 September 2014, Accepted: 15 December 2014Published online: 24 December 2014✉ Correspondence:Jianqiang Li: Phone: +86-25-85560000 (Ext: 3052), Email:************************Hanmei Xu: Phone: +86-25-86185437,Email:139****************VIROLOGICA SINICA 2014, 29 (6): 403-409DOI 10.1007/s12250-014-3508-9I NTRODUCTION Hepatitis B virus belongs to hepadnaviridae , a family of enveloped viruses containing a partially double-stranded and circular DNA genome (G. Birkenmeyer L, 2003). Records show that more than 350 million people world-wide are infected by this virus (Awan A R, et al., 2012). Clinical antiviral therapies currently used for chronic hepatitis B infection often lead to drug tolerance and are unable to eradicate the virus completely (Janssen H L, et al., 2005; Liaw Y F, et al., 2000; Niederau C, et al., 1996). Although an effective prophylactic vaccine has been available for nearly 30 years to date, up to 10%of individuals do not experience a response with anadequate antibody level to hepatitis B surface antigen (Sjogren M H, 2005). Therefore, development of a novel hepatitis B vaccine with a better penetrating and a higher response rate is essential.The HBV envelope consists of three major immuno-genic proteins, known as large (PreS1-PreS2-S), medium (PreS2-S) and small protein (HBsAg), and is the main component of the currently licensed recombinant prophy-lactic vaccine (Hadiji-Abbes N, et al., 2009). Previous studies have suggested that the PreS2-S region has the strongest immunogenicity, which could enhance anti-body response to HBsAg and thus mitigate the chance of non-responsiveness of vaccines against escape variants, making it the most suitable candidate antigen for a novel HBV vaccine (Hadiji-Abbes N, et al., 2009). However, the entire PreS2 region containing 55 amino acids is sen-sitive to protease degradation, which makes large-scale preparation difficult (Langley K E, et al., 1988). The C-terminus of PreS2 containing 26 amino acids (120-145 aa) has been shown to be the most effective immu-Hepatitis B Virus PreS2-S VLP generationVIROLOGICA SINICA404 D ECEMBER 2014 V OLUME 29 I SSUE6nogen for production of antisera and has been proposed to contain a conserved conformational epitope critical for neutralization of infection (Chi S W, et al., 2006). Moreover, PreS2 containing 26 amino acids (120-145 aa) appears more stable than PreS2 containing 55 amino acids. Therefore, a truncated PreS2-S protein comprising 120-145 aa of the PreS2 (26 aa) C-terminus and entire HBsAg was used in the current study.The methylotrophic yeast Hansenula polymorpha (H. polymorpha ), a highly ef fi cient microbial cell factory, has been successfully used in the production of several cur-rent commercial prophylactic HBV vaccines (Gellissen G, et al., 1992; Huang Y, et al., 2007). Here, H. poly-morpha was employed as the host strain to express HBV PresS2-S, leading to a production yield of up to 250 mg per liter. After several steps of puri fi cation, total yield of ~15% with ~99% purity was obtained. Consistent with previous findings, ~22 nm virus-like particles were de-tected using electron microscopy. MATERIALS AND METHODSStrains and reagentsH. polymorpha DL-1 (ATCC 26012) was purchased from ATCC, Geneticin TM (G418) from Invitrogen (Life Technologies, California, USA), the PCR cloning kit from Toyobo (Toyobo, Osaka, Japan) and CsCl from Sigma Aldrich (Sigma Aldrich, Missouri, USA). Eco R I , Not I , Bgl II were purchased from Takara (Takara Bio Inc., Otsu, Japan).Plasmid construction and protein expression The codon-optimized coding sequence of adw2-sub-type HBV PreS2-S antigen (containing 26 aa of PreS2 C-terminus) was amplified from pVAC1010-PreS2-S (plasmid stored in our laboratory). Briefly, the PreS2-S was amplified by KOD plus kit according to the manufacturer’s instructions using primer M26-F: CCGGAATTCGACATGGGTACAGTTAGTC (Eco R I) and primer M-R: ATAGTTTAGCGGCCGCTTAAATGT (Not I). The ampli fi ed coding sequence was subsequent-ly digested with Eco R I, Not I and inserted into the expression vector, pVCH1300. The resultant plasmid was sequenced by GENEWIZ, Inc (Suzhou, China). For yeast transformation, plasmid pV CH1300-PreS2-S was linearized with the restriction endonuclease Bgl II and transformed into H. polymorpha according to the method described by Faber et al. (1994). Positive clones were cultured in BMGY or YPD medium at 220 rpm/min, 37ºC, for 1 day and induced using 1% (w/v) methanol f o r 48 h. Protein puri fi cationHarvested cells was re-suspended in sodium phos-phate buffer (50 mmol/L PB, 150 mmol/L NaCl, pH8.0) and disrupted with a high-pressure homogenizer at 4 ºC (1500 bar, 4 ). The cell debris was removed via centrifugation at 4 ºC, 4,400 g , for 15 min and the super-natant mixed with colloidal silica Aerosil-380 (Evonik Degussa, Essen, Germany ), which was pre-equilibrated in 25 mmol/L sodium phosphate buffer, 0.5 mol/L NaCl, pH 7.2. The suspension was stirred at 4 ºC for 4 h and centrifuged at 4 ºC, 7500 g , for 30 min. The pellet was washed with 25 mmol/L sodium phosphate buffer, pH 7.2, and centrifuged as above. The resultant pellet was re-suspended in 100 mmol/L sodium carbonate-bicarbon-ate buffer (pH 10.8) and incubated at 37 ºC for 4 h. The suspension was centrifuged at 25 ºC, 10,000 rpm, for 1 h. The clari fi ed supernatant was adjusted to pH 8.0 and processed using ion-exchange chromatography (DEAE Sepharose FF, GE Healthcare). Bound PreS2-S was eluted using a salt step (50 mmol/L Tris-HCl buffer, pH 8.0, 0-1 mol/L NaCl). PreS2-S-containing fractions were pooled and further separated via CsCl density gradient centrifu-gation (Beckman, California, USA) at 36,000 rpm and 4 ºC for 16 h.Quantitative measurement of PreS2-S using ELISAThe PreS2-S concentration in the crude extract was determined using a commercially available ELISA kit (Kehua Bio-engineering, Shanghai, China). Purified PreS2-S was employed to establish a standard curve. Crude yeast extract containing PreS2-S was diluted appropriately with 0.1% BSA in PBS (pH 7.2) and ap-plied for ELISA. The resultant absorption values were converted to PreS2-S concentrations using the standard curve established above. All samples were analyzed in triplicate.Silver staining and Western BlotThe protein sample was separated using 12% SDS-PAGE and characterized via silver staining acc ording to the manual of Novex (Life Technologies, California, USA). Western blot analysis was carried out us-ing mouse anti-human HBsAg polyclonal antibody (ABMAX, Beijing, China) and HRP-conjugated goat anti-Mice IgG antibodies (Sigma Aldrich, Missouri,USA ). Brie fly, NC membrane was incubated with pri-mary antibody (1:1000 dilution) in 5% blocking solu-tion at 37°C for 40 min. After washing with TBST, the membrane was incubated with HRP-conjugated second-ary antibody (1:10000 dilution) in 5% blocking buffer at 37°C for 30 min. Signal detection was performed using Western Lightning® Plus-ECL (PerkinElmer. Inc, Massachusetts, USA )Electron Microscopy T h e structure of PreS2-S virus-like particles was con-Xiaowei Xu et al D ECEMBER 2014 V OLUME 29 I SSUE 6 405fi rmed using transmission electron microscopy (HT7700, Hitachi). Purified samples were applied to a 400-mesh copper grid, dried, and stained with 2% tungstophosphor-ic acid before TEM observation. The particle size was measured using Image J software.RESULTSPlasmid constructionThe coding sequence of the adw2-subtype HBV PreS2-S antigen (containing 26 amino acids of the PreS2 C-terminus, 120-145 aa) was optimized according to the codon bias of H. polymorpha (Figure 1A ). The op-timized sequence was ampli fi ed and further cloned into the expression vector, pVCH1300, under the control of the strong methanol-inducing MOX promoter (Figure 1B and 1C ). A geneticin (G418) resistance gene was used as a positive selection marker. The resultant plasmid was veri fi ed using colony PCR and restriction enzyme ana l y-sis (Figure 1D ).Expression of PreS2-SThe plasmid was transformed into H. polymorpha cells using a previously published method (Faber K N, et al.,1994). Multiple copy inserts were screened according to protocols of the Invitrogen manual and further con fi rmed using rea1-time PCR (data not shown). The resultant multiple copy colonies were cultured in YPD medium and induced with methanol at a fi nal concentration of 1% (Figure 2A and 2B ). Induced samples were collected at the indicated time-points and characterized using Western blot (Figure 2C ). After two days of methanol induction, the expression yield was estimated as 250 mg/L culture (2.5 mg/g wet cells).Puri fi cation of HBV PreS2-SHarvested cells was re-suspended in sodium phosphate buffer and disrupted using a high-pressure homogenizer. After clarification via centrifugation at 4,400 g for 15 min, the supernatant containing HBsAg was further clari-fi ed with colloidal silica. The salt steps in the puri fi cation of PreS2-S protein using DEAE Sepharose FF chroma-tography are presented in Figure 3A . The SDS-PAGE pattern (Figure 3B ) showed that the fraction eluted with buffer (50 mmol/L Tris-HCl pH 8.0 plus 500 mmol/L NaCl) mainly contained the monomer and polymer forms of the desired protein and no other evident bands of host protein. PreS2-S-containing fractions were pooled, fur-Figure 1. A: Schematic representation of the truncated PreS2-S. B: Construct of plasmid (pVCH1300-PreS2-S) used to produce Hepatitis B PreS2-S in H. polymorpha. C: The coding sequence of PreS2-S was ampli fi ed byPCR. D: Restriction enzyme analysis of pVCH1300-PreS2-S.Hepatitis B Virus PreS2-S VLP generationVIROLOGICA SINICA406 D ECEMBER 2014 V OLUME 29 I SSUE6ther separated via CsCl density gradient centrifugation, and analyzed via Western Blot. We observed enrichment of PreS2-S mainly in fractions 7-8 (~1.18 g/mL) (Figure 3C ). The final product, with a total yield of ~15% and purity of 99%, was quanti fi ed using ELISA, as described above.Electron Microscopy fi ndingsHBsAg needs to assemble into spherical particles, so-called virus-like particles (VLPs), to initiate the re-quired protective immune response (Cabral G A, et al., 1978). To establish whether PreS2 is able to assemble into VLPs, transmission electron microscopy was per-formed. The hepatitis B PreS2-S antigen derived from H. polymorpha cells organized into 22.6±3.0 (n =32) nm virus-like particles (Figure 4A and 4B ), which were morphologically similar to those isolated from S a ccharomyces cerevisiae and Pichia pastoris . DISCUSSIONHepatitis B virus (HBV) infection is the main cause ofcirrhosis and hepatocellular cancer (Torbenson M, et al., 2002). The currently available recombinant prophylactic vaccine for HBV, either expressed in Chinese hamster ovary (CHO) cells or yeast, can induce effective humoral responses (Chen H, et al., 2012). However, up to 10% of individuals are still unable to experience a response with an adequate protective antibody level to hepatitis B surface antigen (Sjogren M H, 2005). Therefore, there remains an urgent requirement for a novel hepatitis B vaccine with a better penetrating and response rate.The HBV envelope consists of three major immuno-genic proteins, known as large (PreS1-PreS2-S), medi-um (PreS2-S) and small protein (HBsAg), respectively. The PreS2-S region has the strongest immunogenicity, which could accelerate antibody response to HBsAg and thus lower the chance of non-responsiveness of vaccines against escape variants (Hadiji-Abbes N, et al., 2009). Moreover, the PreS2 region contains a conserved polym-erized albumin receptor site involved in virus attachment to human hepatocytes, and anti-PreS2 antibodies often arise earlier than anti-HBsAg antibodies (Kuroda S, et al., 1991; Milich D R, et al., 1985).Figure 2. pVCH1300-PreS2-S transformed H. polymorpha cells growth curves before (A) and after methanolinduction (B). Samples inducing at indicated time points were characterized by Western-Blot (C).Xiaowei Xu et alFigure 3. Purification of HBVPreS2-S from H. polymorphacells. A: Elution profi le of DEAEchromatography. B: Eluate ofDEAE chromatography wascharacterized by Silver Stain-ing. C: Fractions from CsCldensity gradient centrifugationwere analyzed by Western Blot.Figure 4. A: Electron micrograph ofpurified Hepatitis B PreS2-S VLPsderived from H. polymorpha cells. B:Particle size of Hepatitis B PreS2-SVLPs. D ECEMBER 2014 V OLUME 29 I SSUE 6 407Hepatitis B Virus PreS2-S VLP generationVIROLOGICA SINICA408 D ECEMBER 2014 V OLUME 29 I SSUE6However, PreS2-S was comparatively unstable and easily degraded in our previous experiments (data no shown), consistent with other reports (Lian M, et al., 2008). Therefore, a truncated PreS2-S protein compris-ing the C-terminus of PreS2 (120-145 amino acids, 26 aa) and entire HBsAg was used in this study. Vaccines containing PreS2 can compete with HBV and block the route of infection, implying that PreS2 is a potential can-didate component of novel HBV vaccines (Jiang Z G, et al., 2007).The methylotrophic yeast, H. polymorpha , is a highly ef fi cient microbial cell factory (Gellissen G, et al., 1992). Here, a PreS2(26 aa)-S truncated gene was cloned into pVCH1300 and transformed into H. polymorpha DL-1. High dose G418-resistant colonies were screened and confirmed via real-time quantitative PCR. Remarkably, nearly 60 copies of plasmid were integrated into the ge-nome of H. polymorpha DL-1 cells and transferred stably in progeny (data not shown), which possibly contributed to the significant production of foreign proteins. In our experiments, PreS2-S production yield in H. polymorpha cells was up to 250 mg per liter. Additionally, the gen-eration process was more productive and cost-effective. After several steps of puri fi cation, a total yield of ~15% with ~99% purity was obtained, which was significantly higher than that from Pichia pastoris (Hardy E, et al., 2000). As a prerequisite for initiating the protective immune response, the ability to assemble into ~22 nm virus-like particles of PreS2-S antigen derived from H. polymorpha cells was con fi rmed using electron micros-copy (Cabral G A, et al., 1978). To our knowledge, this is the fi rst report of intracellular expression of PreS2-S VLPs in H. polymorpha. Further animal immunization studies with this antigen are required to fully evaluate PreS2-S as a potential vaccine candidate. Efficacy and safety studies will be reported in future publications.ACKNOWLEDGMENTSWe are grateful to our colleagues for sharing their re-sults during scienti fi c meetings and informal discussions. The Jiangsu Simcere Pharmaceutical Group has propri-etary interests in the hepatitis B prophylactic and thera-peutic vaccines described in this study.COMPLIANCE WITH ETHICS GUIDELINES All authors declare no competing interests. This article does not contain studies with human or animal subjects.AUTHOR CONTRIBUTIONSJL and HX designed the experiments. XX, SR, XC, JG, ZX, HH, HS, YG and TZ performed the experiments.XX, SR and JL analyzed data. XX and SR wrote the pa-per. All authors read and approved the fi nal manuscript.REFERENCESA w an A R, Zahoor M Y , Javed M M, Babar M E, Saleem Z. 2012. Expression of pres2/S antigen of hepatitisB virus isolated from Pakistan in yeast cells. Pak. J. Bot., 44: 355–359.C a bral G A, Marciano-Cabral F, Funk G A, Sanchez Y , Hollinger F B, Melnick J L, Dreesman G R. 1978. Cellular and humoral immunity in guinea pigs to two major polypeptides derived from hepatitis B surface antigen. J Gen Virol, 38: 339–350.C h en H, Chuai X, Deng Y, Wen B, Wang W, Xiong S, Ruan L, Tan W. 2012. Optimisation of prime-boost immunization in mice using novel protein-based and recombinant vaccinia (Tian-tan)-based HBV vaccine. PLoS One, 7: e43730.C h i S W, KimD H, Kim J S, Lee M K, Han K H. 2006. Solution conformation of an immunodominant epitope in the hepatitis B virus preS2 surface antigen. Antiviral Res, 72: 207–215.F aber K N, Haima P, Harder W, Veenhuis M, Ab G. 1994. High-ly-ef fi cient electrotransformation of the yeast Hansenula poly-morpha. Curr Genet, 25: 305–310.G . Birkenmeyer L. 2003. Hepatitis B virus: life cycle and mor-phogenesis. In: Ias K M (ed.), Perspectives in Medical Virology, vol. V olume 10. Elsevier, p109–125.G ellissen G, Janowicz Z A, Weydemann U, Melber K, Strasser A W, Hollenberg C P. 1992. High-level expression of foreign genes in Hansenula polymorpha. Biotechnol Adv, 10: 179–189.H adiji-Abbes N, Borchani-Chabchoub I, Triki H, Ellouz R, Gar-gouri A, Mokdad-Gargouri R. 2009. Expression of HBsAg and preS2-S protein in different yeast based system: a comparative analysis. Protein Expr Purif, 66: 131–137.H ardy E, Martinez E, Diago D, Diaz R, Gonzalez D, Herrera L. 2000. Large-scale production of recombinant hepatitis B surface antigen from Pichia pastoris. J Biotechnol, 77: 157–167.H uang Y , Bi J, Zhang Y , Zhou W, Li Y , Zhao L, Su Z. 2007. A highly ef fi cient integrated chromatographic procedure for the puri fi ca-tion of recombinant hepatitis B surface antigen from Hansenula polymorpha. Protein Expr Purif, 56: 301–310.J anssen H L, van Zonneveld M, Senturk H, Zeuzem S, Akarca U S, Cakaloglu Y , Simon C, So T M, Gerken G, de Man R A, Niesters H G, Zondervan P, Hansen B, Schalm S W. 2005. Pe-gylated interferon alfa-2b alone or in combination with lamivu-dine for HBeAg-positive chronic hepatitis B: a randomised trial. Lancet, 365: 123–129.K uroda S, Fujisawa Y , Iino S, Akahane Y , Suzuki H. 1991. Induction of protection level of anti-pre-S2 antibodies in humans immunized with a novel hepatitis B vaccine consisting of M (pre-S2 + S) pro-tein particles (a third generation vaccine). V accine, 9: 163–169.L angley K E, Egan K M, Barendt J M, Parker C G, Bitter G A. 1988. Characterization of purified hepatitis B surface antigen containing pre-S(2) epitopes expressed in Saccharomyces cere-visiae. Gene, 67: 229–245.L ian M, Zhou X, Chen B, Li C, Gu X, Luo M, Zheng X. 2008. Identi fi cation of the critical regions in hepatitis B virus preS re-quired for its stability. J Pept Sci, 14: 307–312.L iaw Y F, Leung N W, Chang T T, Guan R, Tai D I, Ng K Y , Chien R N, Dent J, Roman L, Edmundson S, Lai C L. 2000. Effects of extended lamivudine therapy in Asian patients with chronic hepatitis B. Asia Hepatitis Lamivudine Study Group. Gastroen-terology, 119: 172–180.M ilich D R, Thornton G B, Neurath A R, Kent S B, Michel M L, Tiollais P, Chisari F V . 1985. Enhanced immunogenicity of the pre-S region of hepatitis B surface antigen. Science, 228: 1195–Xiaowei Xu et al D ECEMBER 2014 V OLUME 29 I SSUE 6 4091199. N iederau C, Heintges T, Lange S, Goldmann G, Niederau C M, Mohr L, Haussinger D. 1996. Long-term follow-up of HBeAg-positive patients treated with interferon alfa for chronic hepatitis B. N Engl J Med, 334: 1422–1427.S jogren M H. 2005. Prevention of hepatitis B in nonresponders to initial hepatitis B virus vaccination. Am J Med, 118 Suppl 10A:34S–39S.T orbenson M, Thomas D L. 2002. Occult hepatitis B. Lancet In-fect Dis, 2: 479–486.J iang Z G, Wang Y , Shao X A, Yue Y , Hong X W, Xu L, Xiong S D. 2007. Antibodized gene vaccine harboring to HBV PreS2/S gene can enhance HBV speci fi c CTL response induced by gene immunization. Chinese J Immunol, 963–967. (In Chinese)。
阿尔法拉瓦尔药业线_S用户手册说明书
2. Installation .............................................................................................. 8 2.1. Clearance for dismantling ........................................................................ 8 2.2. Foundations ....................................................................................... 8 2.3. Positioning and slope ............................................................................. 8 2.3.1 Horizontal positioning ............................................................................. 8 2.3.2 Vertical positioning ................................................................................ 8 2.4 Levelling .............................................................................................. 9 2.5 Cleanliness provisions .............................................................................. 9 2.6 Connecting the heat exchanger to the piping system ........................................... 9 2.7 Fitting and piping .................................................................................... 10
泰爱（Telitacicept）注射剂说明书
【药品名称】通用名称：注射用泰它西普商品名称：泰爱英文名称：T elitacicept for Injection汉语拼音：Zhusheyong Taitaxipu【成份】活性成份：泰它西普（重组人B淋巴细胞刺激因子受体－抗体融合蛋白）辅料：盐酸组氨酸、盐酸精氨酸、甘露醇、蔗糖、氢氧化钠【性状】本品应为白色至淡黄色疏松体，复溶后为无色至淡黄色、澄明液体。
【适应症】本品与常规治疗联合，适用于在常规治疗基础上仍具有高疾病活动（例如：抗ds-DNA抗体阳性及低补体、SELENA-SLEDAI评分≥8）的活动性、自身抗体阳性的系统性红斑狼疮（SLE）成年患者。
该适应症是基于一项接受常规治疗仍具有高疾病活动的系统性红斑狼疮成年患者的Ⅱ期临床试验结果给予的附条件批准。
本适应症的完全批准将取决于确证性随机对照临床试验能否证实本品在该患者人群的临床获益。
【规格】80 mg/支。
【用法用量】本品需要在有诊断和治疗系统性红斑狼疮经验的医生指导下使用。
给药方法本品采用皮下注射给药。
注射部位为腹部。
本品为冻干粉，将本品每支（80 mg）用1 ml灭菌注射用水复溶，复溶溶液浓度为每ml含80 mg泰它西普。
复溶时应将灭菌注射用水的水流朝向药瓶的一侧，沿瓶壁缓慢加入，以尽量减少泡沫形成。
复溶期间，将药瓶置于室温条件下，缓慢旋转约60秒，静置至泡沫消退。
切勿摇晃。
待药物溶解后，再次轻轻旋转，药液将会完全混匀。
通常在加入灭菌注射用水后15分钟内完成复溶，但也可能长达30分钟。
复溶后药液为无色至淡黄色、澄明液体。
如观察到可见颗粒，应弃用。
药物溶解后溶液如有小空气泡，不影响使用，但在抽取至无菌注射器后应排除气泡。
本品从复溶到完成皮下注射的总时间不应超过4小时。
剂量本品推荐使用剂量为160 mg/次，每周给药一次。
本品给药期间，经临床医生充分评估患者使用本品的安全耐受性后决定是否需要下调剂量。
如需下调剂量可将每次给药剂量下调为80 mg/次。
益赛普说明书
国内首家“全国连锁专科药房”，也是全国大
型的专科医药连锁企业之一。
坚持“专注于重大慢性疾病用药，持续改善患
者健康”的经营理念。
【益赛普成份】 益赛普主要成份： 每瓶含重组人Ⅱ型肿瘤坏死因子受体－抗体融合 蛋白（rhTNFR:Fc）12.5毫克或25毫克，甘露醇 40毫克，蔗糖10毫克，三羟甲基氨基甲烷1.2毫克。 用1毫升灭菌注射用水溶解。
【益赛普性状】 益赛普为白色冻干粉针剂，加水溶 解后溶液为无色或微带黄色的澄清 透明液体。
【益赛普适应症】 用于治疗：1. 中度及重度活动性类风湿关节炎；2. 18岁及18岁以上成人中度至重度斑块状银屑病；3. 活动性强直性脊柱炎。 【益赛普用法用量】 成人推荐剂量为每次25mg，皮下注射，每周二次。 注射前用1毫升注射用水溶解，溶 解后可冷藏72小时。
【益赛普不良反应】 常见不良反应是注射部位局部反应，包括轻至中度红 斑、搔痒、疼痛和肿胀等，注射部位反应通常发生在 开始治疗的第一个月内，在随后的治疗中发生频率降 低。注射部位反应平均持续3－5天。在临床试验出现 的其它不良事件包括头痛、眩晕、皮疹、咳嗽、腹痛、 淋巴细胞增多白细胞减少、中性粒细胞减少、鼻炎、 发热、关节酸痛、肌肉酸痛、困倦、 面部肿胀、面部过敏、肝功能异常、 肾结石、左肺纤维化等。
百济新特药房资深药师倾力制作千个药品专辑 安全、有效、合理用药之
益赛普篇
商品名：益赛普 厂家：上海中信国健药业有限 公司
【益赛普药品名称】 通用名：注射用重组人Ⅱ型肿瘤坏死因子受体－ 抗体融合蛋白 商品名：益赛普 英文名：Recombinant Human Tumor Necrosis Factor－α ReceptorⅡ:IgG Fc Fusion Protein for Injection 汉语拼音：Zhusheyong Chongzu Ren Erxing Zhongliuhuaisiyinzi Shouti－Kangti Ronghedanbai
薄型子宫内膜小鼠模型的免疫组化鉴定与生育力评估
薄型子宫内膜小鼠模型的免疫组化鉴定与生育力评估张汝月;李瑞娇;张引红;李红霞;郭兴萍【摘要】目的通过免疫组化及生育力评估的手段进一步探讨宫腔注入95％乙醇建立一个稳定的薄型子宫内膜小鼠模型的可行性,为薄型子宫内膜的病理特征及修复机制的相关研究提供理想的动物模型.方法选择78只性成熟且动情期规律的雌性C57BL/6J小鼠作为实验动物,通过宫腔注入95％乙醇损伤子宫内膜建立薄型子宫内膜小鼠模型.将78只实验小鼠随机分为两大组,其中一组30只,又随机分为空白组,对照组和实验组,于造模后第三个动情期将实验小鼠颈椎脱臼法处死,收集子宫样本,免疫组织化学检测细胞角蛋白、波形蛋白、血管内皮生长因子和雌激素受体α的表达以评估子宫内膜细胞的再生情况;另一组48只,又随机分为空白组,对照组,单侧损伤组和双侧损伤组,造模后所有存活小鼠于第三个动情期进行合笼交配,分析薄型子宫内膜对小鼠生育力的影响,评估薄型子宫内膜小鼠模型是否建立成功.结果免疫组织化学结果显示,宫腔注入95％乙醇的实验组小鼠子宫内膜细胞角蛋白、波形蛋白、血管内皮生长因子和雌激素受体α的表达均显著低于空白组和对照组.生育力评估实验显示合笼交配后,空白组和对照组小鼠正常受孕,双侧损伤实验组小鼠未受孕,单侧损伤实验组损伤侧子宫未受孕,未损伤侧子宫正常受孕,且损伤侧子宫的平均怀孕率较未损伤侧显著降低(P<0.05),表明宫腔注入95％乙醇能够损伤小鼠子宫内膜,导致生育力降低.结论本研究成功建立了薄型子宫内膜小鼠模型,该模型可用于薄型子宫内膜修复及修复机制的研究.【期刊名称】《中国比较医学杂志》【年(卷),期】2018(028)012【总页数】8页(P19-26)【关键词】薄型子宫内膜;小鼠模型;免疫组化;生育力评估【作者】张汝月;李瑞娇;张引红;李红霞;郭兴萍【作者单位】山西医科大学基础医学院,太原 030001;山西省生殖科学研究所,出生缺陷与细胞再生山西省重点实验室,太原 030007;山西医科大学实验动物中心,实验动物与人类疾病动物模型山西省重点实验室,太原 030001;山西省生殖科学研究所,出生缺陷与细胞再生山西省重点实验室,太原 030007;山西省生殖科学研究所,出生缺陷与细胞再生山西省重点实验室,太原 030007【正文语种】中文【中图分类】R-33薄型子宫内膜大大影响子宫内膜容受性，导致胚胎着床失败，或不能维持继续妊娠，是生殖医学领域中的难题之一[1]，且薄型子宫内膜作为一个重要的易检测临床指标，受到了广大妇科医生的关注。
益赛普说明书
百济新特药房资深药师倾力制作千个药品专辑 安全、有效、合理用药之
【益赛普适应症】 用于治疗：1. 中度及重度活动性类风湿关节炎；2. 18岁及18岁以上成人中度至重度斑块状银屑病；3. 活动性强直性脊柱炎。 【益赛普用法用量】 成人推荐剂量为每次25mg，皮下注射，每周二次。 注射前用1毫升注射用水溶解，溶 解后可冷藏72小时。
【益赛普不良反应】 常见不良反应是注射部位局部反应，包括轻至中度红 斑、搔痒、疼痛和肿胀等，注射部位反应通常发生在 开始治疗的第一个月内，在随后的治疗中发生频率降 低。注射部位反应平均持续3－5天。在临床试验出现 的其它不良事件包括头痛、眩晕、皮疹、咳嗽、腹痛、 淋巴细胞增多白细胞减少、中性粒细胞减少、鼻炎、 发热、关节酸痛、肌肉酸痛、困倦、 面部肿胀、面部过敏、肝功能异常、 肾结石、左肺纤维化等。
【益赛普成份】 益赛普主要成份： 每瓶含重组人Ⅱ型肿瘤坏死因子受体－抗体融合 蛋白（rhTNFR:Fc）12.5毫克或25毫克，甘露醇 40毫克，蔗糖10毫克，三羟甲基氨基甲烷1.2毫克。 用1毫升灭菌注射用水溶解。
【益赛普性状】 益赛普为白色冻干粉针剂，加水溶 解后溶液为无色或微带黄色的澄清 透明液体。
【益赛普药理作用】
已知TNF是类风湿性关节炎（RA）病理过程中的 一个主要炎性介质，其参与调控的炎症反应可导致 关节的病理改变。益赛普的作用机制为竞争性地与 血中肿瘤坏死因子(TNF)－α结合，阻断它和细胞表 面TNF受体结合，降低其活性。
抗体说明书
PAA121Mu01 Polyclonal Antibody to Stem Cell Factor Receptor (SCFR)Organism Species: Mus musculus (Mouse)Instruction manual FOR IN VITRO USE AND RESEARCH USE ONLYNOT FOR USE IN CLINICAL DIAGNOSTIC PROCEDURES10th Edition (Revised in Jan, 2014) [ PRODUCT INFORMATION ]Immunogen: SCFR, Mouse Clonality: Polyclonal Host: Rabbit Immunoglobulin Type: IgG Purification: Affinity Chromatography. Applications: WB, ICC, IHC-P, IHC-F, ELISA Concentration: 200µg/mLUOM: 100µg[ IMMUNOGEN INFORMATION ]Immunogen: Recombinant SCFR (Tyr343~Pro527) with N-terminal His-Tag expressed in E.coli.Accession No.: RPA121Mu01[ ANTIBODY SPECIFITY ]The antibody is a rabbit polyclonal antibody raised against SCFR. It has been selected for its ability to recognize SCFR in immunohistochemical staining and western blotting.[ APPLICATIONS ]Western blotting: 1:50-400Immunocytochemistry in formalin fixed cells: 1:50-500Immunohistochemistry in formalin fixed frozen section: 1:50-500 Immunohistochemistry in paraffin section: 1:10-100Enzyme-linked Immunosorbent Assay: 1:100-200Optimal working dilutions must be determined by end user.[ CONTENTS ]Form & Buffer: Supplied as solution form in PBS, pH7.4, containing 0.02% NaN 3, 50% glycerol. [ QUALITY CONTROL ]Content: The quality control contains recombinant SCFR (Tyr343~Pro527) disposed in loading buffer.Usage: 10uL per well when 3,3'-Diaminobenzidine(DAB) as the substrate.5uL per well when used in enhanced chemilumescent (ECL). Note: The quality control is specifically manufactured as the positive control. Not used for other purposes.Loading Buffer: 100mM Tris(pH8.8), 2% SDS, 200mM NaCl, 50% glycerol, BPB 0.01%, NaN 3 0.02%. [ STORAGE ]Store at 4o C for frequent use. Stored at -20o C to -80o C in a manual defrost freezer for one year without detectable loss of activity. Avoid repeated freeze-thaw cycles. [ IMAGES ]Used in Western Blot, Sample:Recombinant SCFR, Mouse Used in DAB staining on fromalin fixed paraffin- embedded kidney tissue。
普腾免疫试剂二抗应用说明书
IntroductionApplications ForSecondary AntibodiesIn many biological assays, the target antigen is visualizedindirectly. That is, the secondary antibody, bound to the primary antibody, produces the signal. Since many secondary antibodies can bind to one primary antibody, the result is signal amplification and increased sensitivity.The choice of the appropriate secondary antibody is determined by the specifications (i.e. species, subclass, and fragment type) of the primary antibody and the application. Proteintech ®* offers a variety of secondary antibodies suitable for Western Blotting, ELISA, cellular imaging, and flow cytometry: –Enzyme/Biotin conjugated–Conventional fluorescent conjugatedWestern Blotting ELISAImmunofluorescence Staining ImmunohistochemistryFlow CytometrySECONDARY ANTIBODIES*Proteintech and the Proteintech logo are trademarks of Proteintech Group registered in the US Patent and Trademark Office.Secondary AntibodiesSelecting A SuitableSecondary AntibodyFor Your ApplicationUnderstanding Common Secondary Antibody Nomenclature 1. Host Species Of Primary AntibodyThe secondary antibody has to be raised against the host species of the primary antibody (e.g., anti-mouse, rabbit, rat).2. Class Of Primary AntibodyIn addition to the host specificity, the secondary antibody also has to be specific for the isotype of the primary antibody (e.g., IgG, IgA, IgM).4. Assay RelevanceThe intended application determines the type of conjugate (eg., enzyme orfluorescent conjugated).5. Additional Purification StepsA common purification technique for secondary antibodies is High Affinity Purification, resulting in reduced non-specific binding. In addition, secondary antibodies can go through an additional purification step to reduce potential cross-reactions with other species. Therefore, the secondary antibody solution is passed over different columns containing sera proteins of different species to filter out the non-specific secondary antibody (e.g., for multi-stainings).3. Antibody FragmentsIf the primary antibody is not thewhole antibody but a fragment,the secondary antibody shouldalso be fragment-specific to reducenoise signal (e.g., F(ab’)₂ fragmentsecondary antibodies penetratetissue easier in IHC).Ms Mouse Gt Goat rRb Rabbit Sh SheepRt Rat BV Bovine Hn Human Sw SwineIgG (H+L), heavy and light chain IgG3IgG1 IgG, Fc, receptor bindingIgG2a IgAIgG2bIgMUnderstanding the needs of the primary antibody and assay-dependent requirements are essential for selecting an appropriatesecondary antibody.2Seconday Antibodies3 Enzyme Conjugated/ Biotin Secondary AntibodiesConventional Fluorescent ConjugatedSecondary AntibodiesOPTIMIZE YOUR ANTIBODY EXPERIMENTS WITH PROTEINTECHRequest your free technical guide from .LIVE CHAT TWITTER@proteintechBLOG/news/blog YOUTUBE/ProteintechSupportAvailable 24 hours via Live Chat and 9-5 (CDT) via phone.Email support also available.1 (888) 4PTGLAB (1-888-478-4522) (toll free in USA),or 1(312) 455-8498 (outside USA)1 (312) 455-8408Proteintech Group, Inc.5400 Pearl Street, Suite 300, Rosemont, IL 60018, USA **********************PHONEFAX ADDRESSEMAILPHONE EMAIL FAX ADDRESSEMAIL+44 (161) 8393007+44 (161) 2413103Proteintech Europe, Ltd. 4th Floor,196 Deansgate, Manchester, M3 3WF ***********************************Sales and technical support only.PHONE FAX ADDRESSEMAIL************or 4006-900-926************Wuhan Sanying Biotechnologies D3-3, No.666 Gaoxin Avenue,Wuhan East Lake Hi-tech Development Zone Wuhan, Hubei, P .R.C******************We are ISO 9001 and ISO 13485 accredited.Proteintech GroupUS Head OfficeProteintech EuropeUnited KingdomProteintech EuropeGermanyProteintechChina OfficeCONTACT US。