产ESBLs的肺炎克雷伯菌耐药性及脉冲场凝胶电泳分型研究

目的探讨产超广谱-内酰胺酶(extended-spectrum -lactamases,ESBLs)肺炎克雷伯菌的耐药性状况和分子流行病学特点。方法采用Kerby-Bauer纸片扩散法测定49株临床分离的产ESBLs肺炎克雷伯菌对19种抗菌药物的耐药性;并采用脉冲场凝胶电泳(PFGE)技术对其做同源性分析。结果49株产ESBLs肺炎克雷伯菌对美罗培南和亚胺培南敏感率达100%,对其他17种抗菌药物均表现不同程度耐药;PFGE图谱有6组(A-F),以A型(27株)、B型(10株)、C型(9株)流行为主,49株产ESBLs肺炎克雷伯菌中共有19株和重症监护病房(ICU)相关,占38.8%,流行菌株在各病房之间传播。结论产ESBLs肺炎克雷伯菌呈多重耐药,其对美罗培南和亚胺培南的敏感率为100%,可作为治疗感染的首选药物,其他药物可根据药敏结果选择;浙江大学医学院附属第一医院在2006年2-8月有产ESBLs肺炎克雷伯菌流行,ICU是高发区域,普外科、呼吸内科、综合病房和干部病房中的分离率也较高,需加强检测和控制。     

双重PCR检测急性髓系白血病FLT3-ITD和NPM1基因突变

本研究旨在建立一种同时筛查FLT3-ITD突变和NPM1突变的检测方法.设计2对引物,分别扩增NPM1基因的外显子12和FLT33基因的外显子14、内含子14、外显子15,以覆盖几乎所有已知突变位点.对双重PCR体系的反应程序和引物浓度比例进行优化,将双重PCR产物通过毛细管电泳分离,根据野生型产物和突变型产物的大小差...     

血清苯丙氨酸、酷氨酸的高效毛细管电泳分析

目的 建立和完善用高效毛细管电泳（HPCE）快速测定血清中苯丙氨酸（Phe）和酪氨酸（Tyr）的方法。方法 用HPCE检测7例正常儿童和10例苯丙酮尿症息儿血清中苯丙氨酸和酪氨酸，用苯丙氨酸和酪氨酸标准液测定后绘制校正曲线，并使用高效液相色谱法（HPLC）对部分标本同步测定进行比较。结果 对HPCE实验条件和电泳条件进行优化和选择，最终采用：水浴温度50℃，水浴时间40min，硼酸钠缓冲溶液pH为9．8，分离电压28kV，分离温度15℃。苯丙氨酸迁移时间为6．8min，酪氨酸迁移时间为5．9min;用HPCE检测苯丙酮尿症患儿血清中Phe和Tyr的水平，与HPLC的检测结果差异无统计学意义（P〉0．05）。结论 该方法快速、简便、准确、灵敏度高，为临床检测苯丙酮尿症提供了一种有效的方法。     

毛细管电泳-激光诱导荧光法分离四种神经递质类氨基酸的条件优化

目的探讨毛细管电泳分离神经递质类氨基酸的影响因素。方法采用毛细管电泳法分离4种神经递质类氨基酸的荧光素异硫氰酸酯(FITC-AAs)衍生物,分析电泳缓冲液、表面活性剂以及衍生条件对分离的影响。结果电泳缓冲液的组成、浓度、pH值、表面活性剂以及衍生条件对4种FITC-AA衍生物分离有很大影响,在以75 mm ol/L,pH 9.42硼酸盐含100 mm ol/LSDS为电泳缓冲液,50 mmol/LpH 9.8硼酸盐为衍生缓冲液,4种FITC-AA衍生物20 min内得到很好分离。结论通过条件优化,毛细管电泳-激光诱导荧光技术分离神经递质类氨基酸的分离效果能得到很大提高。     

毛细管电泳技术与分子信标技术结合检测基因

目的：研究毛细管电泳技术和分子信标技术结合进行基因检测，建立一种简便、敏感、特异和快速自动化的基因检测的新方法学。方法：提取胃癌患者的癌组织及癌旁组织DNA，PCR扩增p16基因外显于2，进行琼脂糖电泳检测，SSCP分析，PCR扩增产物与自行设计探针杂交后毛细管电泳检测。结果：有p16外湿子2 PCR扩增产物的癌组织与分子信标探针液相杂交后毛细管电泳仪检测可见检测峰。结论：毛细管电泳技术和分子信标技术结合检测基因产物是在临床及基础研究中可作为一种基因产物检测的实用手段。     

A fluorometric method for determination of C3b inactivator

A simple method for the determination of C3b inactivator in human plasma was investigated. Fluorescent-labeled C3, which was prepared by treatment with methylamine and a fluorescent thiol reagent, N-(dimethylamino-4-methylcoumarinyl)-maleimide, was used as the substrate. Diluted human plasma is incubated with fluorescent C3, reduced with 2-mercaptoethanol, and subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis. The fluorescent cleavage product is then determined by scanning the gels with a fluorospectrophotometer. This method is simple and sensitive enough to determine C3b inactivator activity in 1 μl of human plasma.     

微流控芯片电泳测定血清小而密低密度脂蛋白及临床应用价值

目的建立微流控芯片电泳分离血清小而密低密度脂蛋白(sdLDL)的方法并探讨其临床应用价值。方法利用自制的微流控芯片,选择Tricine缓冲液为电泳缓冲液,SDS作为添加剂,电泳分离经硝基苯并噁二唑-C6-酰基鞘胺醇(NBD C6-ceram-ide)预染的脂蛋白,激光诱导荧光技术检测。结果大而轻低密度脂蛋白(lLDL)、sdLDL、极低密度脂蛋白(VLDL)和高密度脂蛋白(HDL)在3 min内得到有效分离。冠心病(CHD)组,sdLDL检出率显著高于正常对照组(P<0.01)。CHD患者血清经连续3次分析,sdLDL出峰时间和峰面积相对标准差分别为2.18%和2.94%。sdLDL阳性组TG水平增高,HDL-C水平降低,均与阴性组存在显著差异(P<0.01和P<0.05)。CHD患者sdLDL与三酰甘油(TG)、载脂蛋白B(apoB)呈正相关(r=0.82和0.79,P<0.0001)。结论微流控芯片电泳可望成为sdLDL的常规分析手段,用于CHD危险性的评估。     

Changes in atrial natriuretic factor during preload reduction with nitroglycerin in patients with congestive heart failure

Summary. Nine patients with congestive heart failure, New York Heart Association class II-III, were evaluated with right heart catheterization. Plasma atrial natriuretic factor (ANF) was determined in blood samples from the pulmonary artery simultaneously with recordings of right atrial, pulmonary arterial, pulmonary capillary wedge and systemic arterial pressures and heart rate during preload reduction with 0·5 mg nitroglycerin sublingually. Basal plasma ANF levels were higher in patients with congestive heart failure compared to normal controls, and correlated to right atrial, pulmonary arterial, and pulmonary capillary wedge pressures. After nitroglycerin all patients had reductions in right atrial, pulmonary arterial, and pulmonary capillary wedge pressures and a simultaneous decrease in plasma ANF concentrations, reaching lowest values after 10 min. Central pressures and plasma ANF rose to baseline values within 30 min. After nitroglycerin plasma ANF concentrations correlated to pulmonary arterial and pulmonary capillary wedge pressures, while changes in plasma ANF correlated to changes in right atrial and pulmonary arterial pressures. These results provide further evidence that ANF is released by a pressure-sensitive mechanism and demonstrates that ANF secretion in relation to central pressure variations is preserved in patients with congestive heart failure and that the response is rapid.     

Efficacy of novel recombinant fowlpox vaccine against recent Mexican H7N3 highly pathogenic avian influenza virus

Since 2012, H7N3 highly pathogenic avian influenza (HPAI) has produced negative economic and animal welfare impacts on poultry in central Mexico. In the present study, chickens were vaccinated with two different recombinant fowlpox virus vaccines (rFPV-H7/3002 with 2015 H7 hemagglutinin [HA] gene insert, and rFPV-H7/2155 with 2002 H7 HA gene insert), and were then challenged three weeks later with H7N3 HPAI virus (A/chicken/Jalisco/CPA-37905/2015). The rFPV-H7/3002 vaccine conferred 100% protection against mortality and morbidity, and significantly reduced virus shed titers from the respiratory and gastrointestinal tracts. In contrast, 100% of sham and rFPV-H7/2155 vaccinated birds shed virus at higher titers and died within 4?days. Pre- (15/20) and post- (20/20) challenge serum of birds vaccinated with rFPV-H7/3002 had antibodies detectable by hemagglutination inhibition (HI) assay using challenge virus antigen. However, only a few birds (3/20) in the rFPV-H7/2155 vaccinated group had antibodies that reacted against the challenge strain but all birds had antibodies that reacted against the homologous vaccine antigen (A/turkey/Virginia/SEP-66/2002) (20/20). One possible explanation for differences in vaccines efficacy is the antigenic drift between circulating viruses and vaccines. Molecular analysis demonstrated that the Mexican H7N3 strains have continued to rapidly evolve since 2012. In addition, we identified in silico three potential new N-glycosylation sites on the globular head of the H7 HA of A/chicken/Jalisco/CPA-37905/2015 challenge virus, which were absent in 2012 H7N3 outbreak virus. Our results suggested that mutations in the HA antigenic sites including increased glycosylation sites, accumulated in the new circulating Mexican H7 HPAIV strains, altered the recognition of neutralizing antibodies from the older vaccine strain rFPV-H7/2155. Therefore, the protective efficacy of novel rFPV-H7/3002 against recent outbreak Mexican H7N3 HPAIV confirms the importance of frequent updating of vaccines seed strains for long-term effective control of H7 HPAI virus.