化学发光成像法测定人体血清中乙型肝炎病毒

基于对碘苯酚增强的luminol-H2O2-HRP化学发光反应,利用化学发光成像法检测乙肝病毒(HBV)。用该法对人体血清中的乙型肝炎表面抗原、表面抗体、e抗原、e抗体以及核心抗体进行测定,其结果与ELISA法所得结果一致,对表面抗原检测结果为阳性的病人血清测定9次,结果的相对标准偏差为4．2％。     

Expression of Human Hepatitis B Virus Surface Antigen Gene in Transgenic Tobacco

Expression of Human hepatitis B virus surface antigen (HBsAg) gene in plant was reported for the first time. The recombinant plasmid pRoKⅡ-HBsAg was constructed by inserting HBsAg gene into the downstream of CaMV 35S promoter of binary vector pRoKⅡ and then introduced into Agrobacterium tumefaciens LBA4404. The kanamycin-resistant plants were obtained by Agrobacterium-mediated transformation system. It was shown that HBsAg gene was expressed in transgenic tobacco plants and their progenies by ELISA. The spherical particles of ψ 22 nm in the leaf extract of trangenic tobacco were observed by immunosorbent electron microscopy.     

高效液相色谱-串联质谱法同时测定化妆品中的两种补骨脂素

建立了高效液相色谱-串联质谱（HPLC-MS/MS）测定化妆品中 8-甲氧基补骨脂素和 5-甲氧基补骨脂素含量的方法。以甲醇提取样品,以甲醇和水为流动相,经HPLC分离后以多级反应监测(MRM)测定。两种补骨脂素在2.0~100.0 μg/L范围内线性良好,检出限均为0.10 mg/kg,回收率分别在96.88%~101.81%和94.25%~101.26%之间,相对标准偏差分别在0.40%~6.02%和0.76%~4.26%之间。     

Isolation and Purification of Recombinant HBsAg of Human Hepatitis B Virus from Silkworm Larvae

Recombinant HBsAg coded by preS1-preS2-S regions of hepatitis B virus was expressed in Bombyx mori silkworm larvae. Recombinant HBsAg was expressed (30–40 μg/mL, 0.1% of the total amount of extracted protein) by larvae infected with recombinant baculovirus rBmNPV-Hep-preS1-S containing cDNA of HBsAg strictly by the polyhedrin gene promoter. Recombinant HBsAg consisting of a polypeptide of molecular weight ∼36 kDa (p36) was purified by gel filtration and affinity chromatography to 92% purity. __________ Translated from Khimiya Prirodnykh Soedinenii, No. 5, pp. 477–480, September–October, 2005.     

基于毛细管液滴技术的血液乙型肝炎病毒DNA提取

基于毛细管液滴技术,建立了提取血液中乙型肝炎病毒(HBV)DNA的方法.方法的基本原理是向聚四氟乙烯毛细管中连续引人油相和水相溶液时,由于表面张力作用,可以形成稳定的油包水型液滴.依次引人含有不同试样的液滴,在毛细管中完成进样、DNA结合、洗涤以及洗脱等过程.实验表明,采用蛋白酶K裂解和提高洗脱温度有利于提高DNA回收...     

分子信标用于不对称PCR检测乙型肝炎病毒

为了缓解竞争杂交对分子信标检测的影响，将分子信标与不对称聚合酶链式反应(PCR)联用进行血清中乙型肝炎病毒(HBV)的检测。并采用更为直接的荧光方法，将不对称PCR扩增中的引物浓度比确定为5：1，该结果与电泳方法得到的结果有所不同。文中结合对称及不对称PCR过程中荧光强度的变化情况，对其原因进行了详细的探讨。分子信标与不对称PCR联用，可以专一地检测出血清中HBV的存在，与分子信标／对称PCR相比．其检测效果明显增强。     

国内外酶免疫试剂检测乙肝病毒标志物的比较

报道了国产与美国Ａｂｂｏｔｔ公司生产的ＨＢＶＥＬＩＳＡ诊断试剂盒,对３０例来自昆明医学院传染科门诊及住院部标本同时进行ＨＢｓＡｇ、抗－ＨＢｓ、ＨＢｅＡｇ、抗－ＨＢｅ及抗－ＨＢｃＩｇＭ检测,对两试剂阳性符合率进行了比较研究,并采用滴定ＨＢｓＡｇ及抗－ＨＢｓ阳性血清的方法,表明两试剂敏感性极为相似,而国产试剂对抗－ＨＢｃＩｇＭ阳性检出率大大提高于Ａｂｂｏｔｔ试剂。     

血清样品中乙肝病毒的DNA电化学传感器检测

利用自组装单分子膜技术，将巯己基修饰的具有乙肝病毒(HBV)DNA序列特异性的单链DNA探针固定在金电极表面，制得DNA电化学传感器；以电活性的Hoechst 33258为指示剂，考察了该传感器对血清样品中乙肝病毒DNA的响应；探讨DNA电化学传感器在临床检测中的应用；将传感器法与荧光聚合酶链反应(PCR)法进行对比，两者的分析结果具有一致性。     

Autologous Materials in Regenerative Dentistry: Harvested Bone,Platelet Concentrates and Dentin Derivates

The jawbone is a peculiar type of bone tissue, unique for its histological, anatomical and physiological characteristics. Therefore, a defect in the maxilla or in the mandible, because of pathological sequelae is difficult to prevent and to restore. Several biomaterials have been and are currently being developed to respond to the demands of regenerative medicine. A specific group of biomaterials used in regenerative dentistry is represented by the autologous materials. Platelet concentrates harvested bone and dentin derivates are indeed used in an attempt to minimise the alveolar resorption or in vertical ridge augmentation procedures or in sinus lift interventions. The aim of this review is to examine the properties of the above-listed materials, to compare them and to indicate eventual clinical applications.     

8-Methoxypsoralen and Ultraviolet A Radiation Activate the Human Elastin Promoter in Transgenic Mice: In vivo and in vitro Evidence for Gene Induction

Treatment of skin diseases with the combination of 8-methoxypsoralen and ultraviolet A radiation (PUVA) results in clinical alterations in treated skin that resemble those observed in chronically photodamaged skin. The PUVA-treated patients develop nonmelanoma skin cancers, pigmentary alterations and wrinkling characteristic of sun-induced changes. The major alteration in the dermis of sun-damaged skin is the deposition of abnormal elastic fibers, termed solar elastosis. Up-regulation of elastin promoter activity in dermal fibroblasts explains the excess elastic tissue but not the reason for the aberrant morphology of the elastotic material. In order to study photoaging in an experimental system, we utilized a transgenic mouse line that expresses the human elastin promoter/chloramphenicol acetyltransferase construct in a tissue-specific and developmentally regulated manner. Although UVB radiation has been demonstrated to increase promoter activity in vitro, UVA fails to demonstrate a similar effect at the doses utilized. In this study, we demonstrate the ability of PUVA treatment to up-re-gulate elastin promoter activity both in vitro and in vivo. These data help to explain the development of photoaging in sun-protected PUVA-treated skin. We attribute the up-regulation of elastin promoter activity in response to PUVA to the formation of DNA photoadducts, which do not occur in response to UVA radiation alone.     

琼脂糖凝胶蛋白芯片片基的制备及应用

建立了一种制备琼脂糖凝胶修饰玻片的方法。以乙肝表面抗原为检测指标,比较了琼脂糖凝胶片基和常规的醛基片基固定抗体后的检测灵敏度,发现琼脂糖凝胶修饰后可以明显增加检测的荧光强度,并在10ng/L～100μg/L之间具有良好的线性关系。在此片基同时固定了抗HBsAg和抗HBeAg抗体,并以牛血清白蛋白和生物素标记抗体为阴性和阳性对照点,采用夹心式免疫反应法检测乙肝病毒的两种抗原,取得良好效果。     

4253例乙型肝炎病毒血清标志物定性结果模式分析

为适应乙肝两对半模式的变化,更准确地进行乙肝诊断,对4 253例乙型肝炎病毒血清标志物(HBVM)的结果模式特征进行了分析。采用ELISA法检测乙型肝炎病毒血清标志物(1-HBsAg、2-抗HBs、3-HBeAg、4-抗HBe、5-抗HBc)。结果表明,ELISA检测HBVM共得到18种模式,其中"全部阴性"占18.3%;"1、4、5阳性"比例最高占23.5%;"1、3、5阳性"占18.1%;"2阳性"占15.1%;"1、5阳性"占12.1%;"2、4、5阳性"占4.3%;"4、5阳性"占2.1%;"2、5阳性"占2.6%;"5阳性"占2.1%;其它特殊模式所占比例相对较低。     

商品化血糖仪用于乙型肝炎病毒的便携式体外分子诊断

为解决当前乙型肝炎病毒(HBV)基因检测技术在一定程度上存在成本高、操作繁琐、误诊率高等问题,在此提出了一种便携式生物传感平台用于HBV基因的超灵敏检测,可检出低至2 copies/μL的HBV基因.首次通过设计针对HBV检测序列的环介导等温扩增(LAMP)反应,实现了在40 min之内对HBV基因型单拷贝基因的检测....     

乙型肝炎患者血清中SOD与LPO含量分析

为了研究乙型肝炎患者血清中超氧化物歧化酶（SOD）活性水平、过氧化脂质（LPO）含量及其变化规律,应用黄嘌吟氧化酶法和硫代巴比妥酸法测定了53例乙肝患者血清中SOD与LPO含量,其中急性乙型肝炎（急肝）35例,慢性活动性乙型肝炎（慢活肝）18例。结果表明,乙型肝炎患者（包括急肝、慢活肝）血清中SOD的含量高于正常对照组（P＜0．05）,而LPO的含量也高于对照组（P＜0．01）,具有显著性差异,提示乙型肝炎患者体内存在着自由基紊乱。     

硒抑制人癌细胞株和人B细胞的EB病毒效应

应用间接免疫荧光法和显微荧光分光光度技术,研究了试管内无细胞毒性浓度的亚硒酸钠对来源于伯基特淋巴瘤的Ｒａｊｉ细胞株,Ｅｐｓｔｅｉｎ－Ｂａｒｒ病毒抗原表达的影响以及抑制ＥＢＶ转化人脐血Ｂ淋巴细胞的作用。     

Immune Responses to a Dicistronic Plasmid Expressing HBsAg of Hepatitis B Virus and Interferon-γ

DNA vaccines encoding a viral protein have been shown to induce antiviral immune responses and provide protection against subsequent viral challenge. The present article deals with the efficacy of a DNA vaccine greatly improved by the simultaneous expression of HBsAg and interferon-γ gene. We constructed a dual expression vector pHIN encoding the HBsAg of Hepatitis B virus and murine IFN-γ which are connected with Internal Ribosome Entry Site(IRES). Mice inmunized with this dual expression DNA vaccine exhibited the enhancement of cellular immune response and increased the production of anti-HBV surface antibody, compared with the mice of single gene expression control. Taken together, these results demonstrate that the application of a cytokine gene in a DNA vaccine formulation as an adjuvant can improve its immunigenicity.     

由质谱分析数据建立乙肝病人与健康人的分类模型

建立运用表面加强激光解吸电离-飞行时间质谱获取乙肝病人和健康人血清蛋白指纹图谱数据,并用偏最小二乘(PLS)变量筛选法建立乙型肝炎(HBV)病人和健康人的分类模型,最终得到分类模型的交叉检验相关系数达0.97以上,判别准确率显著提高.对模型进行分析,找出对乙型肝炎病人和健康人的差异有重要影响的因素或变量.这些变量为某些质荷比区间内特定蛋白的峰强度值,反映这些质荷比区间内蛋白量的增加或减少,与乙肝病的形成有密切关系,可作为重要的生物标志物,进一步加以研究.本研究采用所得模型的拟合值等一些信息来做分类图,能较好地表达回归模型的分类效果.     

Application of Single—labelled Probe—primer in PCR Amplification to the Detection of Hepatitis B Virus DNA

A new method based on the incorporation of a single-lablled probe-primer into polymerase chain reaction(PCR) for the detection of PCR-amplified DNA in a closed system is reported.The probeprimerc consists of a specific probe sequence on the 5‘‘‘‘‘‘‘‘-end and a primer sequence on the 3‘‘‘‘‘‘‘‘-end.A flurophore is located at the 5‘‘‘‘‘‘‘‘end.The primeR-quencher is an oligonucleotide,which is complementary to the probe sequence of probe-primer and labelled with a quencher at the 3‘‘‘‘‘‘‘‘-end.In the duplex formed by probe-primer and primer-quencher.the fluorophore and quencher are kept in close proximity to each other.Therefore the fluorescence is quenched.During PCR amplificatio,the specific probe sequence of probeprimer binds to its complement within the same strand of DNA,and is cleaved by Taq DNA polymerase,resulting in the restoration of fluorescence.This system has the same energy transfer mechanism as molecular beacons,and a good quenching effciency can be ensured.Following optimization of PCR conditions,this method was used to detect hepatitis b virus(HBV) dna in patient sera.This technology eliminates the risk of carry-over contamination,simplifies the amplification assay and opens up new possibilities for the real-time detection of the amplified DNA.     

Immune Responses to a Dicistronic Plasmid Expressing HBsAg of Hepatitis B Virus and Interferon-γ

DNA vaccines encoding a viral protein have been shown to induce antiviral immune responses and provide protection against subsequent viral challenge.The present article deals with the efficacy of a DNA vaccine greatly improved by the simultaneous expression of HBsAg and interferon-γ gene.We constructed a dual expression vector pHIN encoding the HBsAg of Hepatitis B virus and murine IFN-γ which are connected with Internal Ribosome Entry Site(IRES).Mice immunized with this dual expression DNA vaccine exhibited the enhancement of cellular immune response and increased the production of anti-HBV surface antibody,compared with the mice of single gene expression control.Taken together,these results demonstrate that the application of a cytokine gene in a DNA vaccine formulation as an adjuvant can improve its immunigenicity.