重组纤维连接蛋白EDA-EDB融合蛋白的分子克隆

目的构建重组pET28a-EDA-EDB质粒,制备重组纤维连接蛋白EDA-EDB融合蛋白。方法采用基因重组技术,将EDA、EDB基因片段连接,再将该重组基因插入pET28a表达载体,表达重组融合EDA-EDB蛋白后,利用6×His/Ni-NTA纯化系统进行纯化,免疫印迹法检测。结果成功连接EDA、EDB基因片段,重组pET28a-EDA-EDB质粒;在大肠杆菌BL21(DE3)中高度表达重组蛋白,获得较纯的重组蛋白,免疫印迹法证实为FN的组分。结论EDA-EDB重组蛋白能采用pET系统在大肠杆菌中表达,并可通过6×His/Ni-NTA纯化系统进行纯化而获得免疫原。     

Giving until it hurts: prisoners are not the answer to the national organ shortage

This Note argues that prisoners, whether executed or living, should not become organ donors. The introduction acknowledges the shortage of transplantable organs in the United States and the steps that have been taken to ameliorate the crisis. Part I discusses the procurement of organs from executed prisoners, beginning with a brief examination of China, a country where this type of procurement is routinely practiced. Part I also examines organ procurement legislation pertaining to executed prisoners. Finally, Part I asserts the reasons that prisoners should not become donors, including the dead donor rule, the ban against physicians as executioners, the Oath of Hippocrates, the risk of transmissible diseases, and the negative perception that would result if organ procurement was tied to executions. Part II of this Note discusses prisoners donating their organs in return for mitigated sentences. Part II then argues that this practice should not be adopted because of the lack of informed consent and voluntary choice. Finally, Part III of this Note introduces potential solutions to the possibility of maintaining a voluntary system, moving to a presumed consent system, and using financial inducements to create a larger supply of transplantable organs.     

A kinetic model describing the pharmacokinetics of ethyl glucuronide in humans

The glucuronide conjugation is a minor pathway of ethanol metabolism. The determination of ethyl glucuronide (EG) in serum and urine has gained importance in forensic and other legal decisions. To prospectively calculate the serum concentration of this non-oxidative ethanol metabolite, the computer program developed includes a parameter fitting routine. Multiple ethanol doses can be handled.The mathematical modeling was based on the following assumptions and simplifications, respectively. A single enzyme system is responsible for ethanol conjugation at one distinct site; the distribution of EG into the systemic circulation is delayed; the elimination of EG follows first-order kinetics.The concentration of EG was calculated using three kinetic parameters: a rate constant for the first-order formation of EG from serum ethanol, a transfer constant for an obstructed transfer of EG from the formation site (FS) into the central compartment (CC) and an exponential elimination constant.The program was applied to the data collected from 21 drinking experiments. The fitting algorithm optimized the three kinetic parameters, until the sum of concentration error squares of the data points was minimized. The means+/-standard deviation of the rate constant for the first-order formation of EG from serum ethanol was 0.0011+/-0.0006 h(-1), the transfer constant for an obstructed transfer of EG from the FS into the CC was 0.43+/-0.1996 h(-1) and the exponential elimination constant was 3.0+/-1.45 h(-1).Using the range of these parameters, it is now possible to calculate minimum and maximum serum concentrations of EG based on ethanol doses and drinking times. The comparison of calculated and measured concentrations can prove the plausibility of an alleged ethanol consumption. This can be crucial when the serum ethanol concentration (SEC) itself is not meaningful.     

An unusual case of driving under the influence of enflurane

An unusual case of driving under the influence of the volatile anaesthetic enflurane is reported. A markedly affected anaesthetist was sniffing at an enflurane-moistened handkerchief before he crashed into a lorry at a red traffic light. In the blood sample enflurane (2.92 mg/l) as well as diclofenac (0.28 mg/l) and 4-aminophenazone (24.4 mg/l) were found. The way of driving and the accident and the deficiency symptoms could be explained by the central suppressing effects of enflurane. The physician was considered as impaired and not suitable to drive at the time of the incidence.     

Quantitation using GC-TOF-MS: example of bromazepam

Time-of-flight mass spectrometry (TOF-MS) offers new perspectives for forensic toxicology. Qualitative and quantitative analyses of a mixture of three selected benzodiazepines (diazepam, nordazepam and bromazepam) were used to compare gas chromatography (GC-TOF-MS, quadrupole GC-MS, GC-ECD) and liquid chromatography (HPLC-DAD) data. Method validation parameters like LOD, LOQ, S/N-ratios reflect the capabilities of GC-TOF-MS. Five-point calibrations for bromazepam in human peripheral blood (50, 100, 160, 200, 300 ng/ml) using medazepam as internal standard (1000 ng/ml) were performed. The calibrations using GC-TOF-MS (using the fragments of m/z 236 and 288), GC-ECD (dual system) and HPLC-DAD (at 235 nm) all showed correlation coefficients close or superior to 0.99. Quadrupole GC-MS data was not used in the comparison of extracted samples due to the low sensitivity in the full scan mode. Two analyses of real cases concerning bromazepam are presented. In the first case, the presence or absence of bromazepam could not be established with both HPLC-DAD and GC-ECD due to background signals. The extracted ion chromatograms and spectrum traces after the analysis with the GC-TOF-MS could clearly excluded the presence of bromazepam. The second case illustrates the quantitation of bromazepam, where both HPLC-DAD and GC-ECD were unable to give satisfactory results, again due to interfering background signals. The analyses performed on the GC-TOF-MS-system demonstrated high sensitivity and also high selectivity due to the high quality of mass spectra obtained. The advantages of GC-TOF-MS make it a promising analytical technique for forensic toxicology.     

Analysis of dimethylamphetamine and its metabolites in human urine by liquid chromatography-electrospray ionization-mass spectrometry with direct sample injection

An analytical method to identify and determine dimethylamphetamine (DMA) and its metabolites in human urine was developed with liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS) involving the direct injection of a urine sample. The urine samples were directly injected by using a gel permeation column, whose stationary phase was polyvinyl alcohol with a small amount of a carboxyl group, so DMA and its metabolites were analyzed rapidly and simply without pretreatment such as extraction, concentration and derivatization. DMA and its metabolites were identified in drug-free human urine spiked with 1 microg of DMA, dimethylamphetamine N-oxide (DMANO) and methamphetamine (MA), and 3 microg of amphetamine (AM) in 1 ml of urine under the full-scan mode. Under the selected ion monitoring (SIM) mode, the limits of detection (signal-to-noise ratio=5) for DMA, DMANO, MA and AM were 20, 20, 20 and 60 ng in 1 ml of urine, respectively. This method was applied to the identification and determination of DMA and its metabolites in urine samples of 10 DMA abusers. The concentrations of DMANO were higher than those of unchanged DMA in all urine samples; thus, DMANO is considered to be a useful metabolite as an indicator to prove DMA intake.     

Use of the "SMITEST" PSA card to identify the presence of prostate-specific antigen in semen and male urine

To determine whether the prostate-specific antigen (PSA) could be identified in semen using the "SMITEST" PSA immunochromatographic membrane test card, we examined semen and other body fluids, including urine. Although PSA activity was detected in semen with high sensitivity using the "SMITEST" PSA card, it was also detected in adult male urine. However, the lower detectable limit in the urine was 1000-fold lower than that in semen. The concentration of PSA in adult male urine was found to be 800 ng/ml using the card. PSA activity usually can be detected in urine of individuals over 14 years old and it has been detected in urine from children as young as 11 years old. Therefore, the appearance of PSA in urine may occur anytime between the age of 12 and 14 years. To determine the stability of PSA activity in urine, dried samples of urine on filter paper were kept at room temperature for up to 3 years. Although the immunoreactive line showing PSA activity became weak after storage, it was still detectable, but faint, after 3 years. In addition, PSA activity was not detected in male serum or saliva and in the urine from human females, male cats or male dogs using the PSA card. We conclude that the PSA card is useful for identification of PSA in both semen and adult male urine.     

美国与东亚大国的关系--东南亚的视角

本文评述了美国与东亚大国———中国、日本、俄罗斯、印度及东盟的关系,以及这些关系对东亚的影响,并着重分析了中国与东亚大国的关系。作者认为,中美关系肯定是未来东亚地区最重要的战略关系。中国的崛起促使创立东亚地区机制成为必要。多边合作是解决和克服双边低效的最直接的战略,因此,创立东亚共同体的想法具有很重要的意义。     

印尼民主化进程、中国印尼关系及国际形势评析

应暨南大学的邀请,2006年5月27-30日,印尼前国家领导人———印尼人民协商会议(MPR)主席、印尼国民使命党前任总主席、穆罕默迪亚前任总主席、现印尼国民使命党荣誉主席、印尼知名学者阿敏.赖斯教授(Prof.Dr.HM Amien Rais)对暨南大学进行了为期3天的学术访问,并被聘请为暨南大学客座教授。期间,阿敏.赖斯教授做了两场学术报告,分别就印尼民主化进程、中国印尼关系、国际形势等诸多问题发表了看法。参加学术报告会的有暨南大学东南亚研究所暨国际关系学系师生、印尼留学生、广东外语外贸大学印尼语专业师生和广州市、中山市、珠海市的印尼归侨代表等130人,东南亚研究所所长曹云华教授主持了学术报告会。现对其主要观点和看法进行整理,以飨广大