VEGF189基因合成、原核表达及鉴定

目的 获得编码VEGF189蛋白的基因,并通过原核表达系统表达VEGF189融合蛋白.方法 应用SOE PCR技术合成VEGF189基因,克隆入pET-32a(+)原核表达载体,转化E coli BL21(DE3)感受态细胞,IPTG诱导表达,进行SDS-PAGE和Western blot检测,表达产物经HisTrapTM HP亲和柱层析纯化,测定蛋白浓度.结果 DNA测序分析表明,合成的VEGF189基因与文献报道相一致.在大肠杆菌BL21(DE3)中表达的Trx-VEGF189融合蛋白,以包涵体和可溶2种形式存在.表达产物可被兔抗鼠VEGF多抗特异识别.经HisTrapTM HP亲和柱纯化,获得纯度达85%的融合蛋白.结论 成功合成VEGF189基因,并在原核表达系统中获得高表达,为研制高效抗肿瘤的VEGF抗体和疫苗奠定了前期基础.     

Effects of fluvastatin on biliary lipids in subjects with an elevated cholesterol saturation index

OBJECTIVE: 3-Hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitors have been suggested as agents to reduce the biliary cholesterol saturation index (CSI) in duodenal bile and therefore might be supportive in primary or secondary prevention of gallstones. However, the efficiency of the therapy seems to depend on both the HMG-CoA reductase inhibitor used and the study population selected. METHODS: We therefore investigated the effect of a high-dose application of fluvastatin on biliary lipid composition in 21 subjects exhibiting mild hypercholesterolaemia and a history of current gallstones or cholecystectomy due to gallstone disease. Subjects were treated either with 40 mg fluvastatin twice per day over a 3-month period (n = 14) or with placebo (n = 7). Bile samples were aspirated during endoscopy after intravenous ceruletid stimulation before and after therapy. RESULTS: Both groups were comparable in CSI (mean +/- SD) at baseline (1.78 +/- 0.2 placebo vs. 1.97 +/- 0.4 verum). CSI significantly decreased in the verum group to 1.45 +/- 0.4 (P = 0.003) mainly due to increased phospholipid levels, whereas no difference was observed in the placebo group (1.85 +/- 0.7, n.s.). In addition, the verum group exhibited a significant reduction of hydrophobic deoxycholic acid, which has been reported to induce cholesterol crystal precipitation, and an increase of hydrophilic cholic acid. CONCLUSION: Fluvastatin might decrease the risk of cholesterol gallstone formation in patients with elevated biliary CSI during long-term treatment by reduction of biliary cholesterol saturation and percentage change in deoxycholic acid content.     

系列染色法分析人－（人－鼠）杂交瘤C8的染色体

Ｃ８杂交瘤分泌人的ＩｇＭ（λ）单克隆抗体，是具有肾综合征出血热病毒抗原内影像作用的Ａｂ２（β）我们用常规Ｇｉｅｍｓａ染色、Ｇ－分带和Ｃ－分带的系列染色法对Ｃ８杂交瘤细胞的染色体进行分析，发现除了小鼠染色体外，还含有人的１，２，５，７，９，１２，１６，１７，１８，１９，２０，２１和２２号染色体及大量标记染色体，人染色体中以较短的染色体出现频率较高。用系列染色法分析人－鼠杂交瘤中的染色体比单独用Ｇ分带法更易得到可靠的结论。     

含现症感染条带血清抗体检测幽门螺杆菌现症感染的多中心临床研究

目的 评价应用Genelabs安速TM试剂盒的现症感染条带(CIM)检测诊断幽门螺杆菌(Hp)现症感染的可行性.方法 采用全国性多中心开放性平行对照临床研究,研究对象为未接受过Hp根除治疗的需要接受Hp感染检测患者.共有7个中心参加本项研究,2007年1至12月纳入300例符合入选标准的被检测者,平均年龄(41±16)岁,其中男128例,女172例.对所有患者均采用13C-尿素呼气试验及Genelabs安速TMHp快速检测试剂盒CIM检测Hp感染情况.13C-尿素呼气试验阳性即认为存在Hp现症感染,13C-尿素呼气试验阴性判断为Hp阴性.结果 经13C-尿素呼气试验检测阳性131例,阴性169例.Genelabs安速TM试剂盒血清抗体检测Hp现症感染的敏感度为87.8%(115/131),特异度为89.9%(152/169),阳性预测值为87.1%(115/132),阴性预测值为90.5%(152/168),准确度为89.0%(267/300).结论 Genelabs安速TMHp快速检测试剂盒可以用于Hp现症感染的筛查,对于既往未接受过根除治疗的患者,其检测阳性可以认为存在Hp现症感染.

Abstract：

Objective To evaluate the performance of the ASSURETM Helwobacter pylori (H.pylori)rapid test[current infection marker(CIM)kit,Genelabs Diagnostics]in detecting the current infection marker CIM for the diagnosis of H.pylori in adult patients.Methods For this multicenter and controlled clinical trial,a total of 300 patients with an average age of(41±16)years old were recruited from 7 participating hospitals.There were 128 men and 172 women.never received any H.pylori eradication therapy.Each subject received a 13C-urea breath test(13C-UBT)and a CIM test.Such performance parameters as sensitivity,specificity,positive and negative predictive values and accuracy were determined by 13C-UBT for the diagnosis of current H.pylori infection.Results According to the gold standard,there were 131 positive and 169 negative subjects.The accuracy rate,sensitivity,specificity,positive and negative predictive values of CIM test were 89.0%(267/300),87.8%(115/131),89.9%(152/169),87.1%(115/132)and 90.5%(152/168)respectively.Conclusion As a simple,rapid,accurate and affordable assay.CIM test may be useful for a non-invasive diagnosis of H.pylori infection in cases without eradication therapy.     

SARS-CoV GDH株N蛋白全长基因克隆及其可溶性表达

目的构建SARS冠状病毒N蛋白的原核表达质粒,诱导重组蛋白表达并纯化,鉴定其抗原性。方法以我国SARS冠状病毒GDH株总RNA为模板,采用RT-PCR技术扩增N蛋白的全长基因,TA克隆后测序。构建pET-23d的N基因表达载体,用IPTG诱导目的蛋白表达,利用硫酸铵沉淀、分子筛层析及离子交换层析纯化重组蛋白,免疫印迹鉴定重组蛋白。结果RT-PCR扩增出1269bpSARS冠状病毒N蛋白的基因片段,其序列分析结果与SARS-CoVGD01、BJ01株的同源性为99.92%;该基因在大肠杆菌表达系统中高效表达,占可溶性蛋白的33.57%,表达产物为非融合的可溶性蛋白,Westernblot结果显示重组N蛋白具有良好的抗原性;纯化后重组N蛋白纯度为92.9%。结论成功构建了SARS冠状病毒N蛋白的重组表达质粒,并在大肠杆菌中以非融合蛋白的形式得到高效的可溶性表达,为SRAS的诊断和疫苗的研制奠定了基础。     

GD2抗体达妥昔单抗β治疗神经母细胞瘤的临床应用专家共识（2021年版）

神经母细胞瘤(neuroblastoma,NB)是儿童最常见的颅外实体肿瘤,临床表现及预后具有高度异质性。低危患儿预后较好,高危患儿即使接受化疗、放疗、手术、造血干细胞移植等多种方法的综合治疗,长期存活率仍不足50%。探索潜在的治疗靶点对于提高高危患者的生存率具有重要意义。双唾液酸神经节苷脂抗原GD2在NB细胞高表达,达妥昔单抗β可与NB细胞膜表面过表达的GD2特定靶点结合,触发抗体依赖性细胞介导的细胞毒性作用和补体依赖的细胞毒性效应,通过双重免疫机制而发挥抗肿瘤作用。通过总结国外多项关键临床研究,并结合在海南和天津医疗先行区的上市前初步应用经验,我们对达妥昔单抗β的安全性、有效性、适用人群及使用方法进行总结和推荐,提出本共识,旨在为临床医师提供指导与帮助。     

亲和层析纯化重组人源性抗HBsAg Fab抗体

目的：纯化酵母发酵产生的重组人源性抗HBsAg Fab抗体，建立稳定、适于生产应用的纯化工艺。方法：以原核表达的重组人源性抗HBs scFv免疫BALB／c小鼠，经杂交瘤技术制备大量分泌mAb的杂交瘤细胞株14F7。将该细胞株注入小鼠腹腔制备mAb腹水，经辛酸沉淀纯化后，制备亲和层析柱。纯化酵母发酵产生的重组人源性抗HBsAg Fab抗体。结果：用抗scFv mAb亲和层析柱纯化的重组人源性抗HBsAg Fab的纯度可达95％以上，回收率可达70％～85％。结论：人源性抗HBs scFv制备mAb作为亲和层析的介质，可有效地从酵母发酵上清中纯化重组人源性抗HBsAg Fab，适用于大规模生产重组人源性抗HBsAg Fab。     

Development and optimization of an antibody array method for potential cancer biomarker detection

Biomarkers play an important role in the detection at an early stage of pancreatic cancer. The aim of the present study was to optimize the conditions of antibody arrays for detecting Hippocalcin-like 1 (HPCAL1), phosphatidylethanolamine binding protein 1 (PEBP1), lectin galactoside-binding soluble 7 (LGALS7), and serpin peptidase inhibitor clade E member 2 (SERPINE2) as biomarkers for pancreatic cancer detection in a single assay and to investigate antibodies’ specificity and cross-reactivity. Capture anti...