Evaluation of Skin Surface as an Alternative Source of Reference DNA Samples: A Pilot Study

An acceptable area for collecting DNA reference sample is a part of the forensic DNA analysis development. The aim of this study was to evaluate skin surface cells (SSC) as an alternate source of reference DNA sample. From each volunteer (n = 10), six samples from skin surface areas (forearm and fingertips) and two traditional samples (blood and buccal cells) were collected. Genomic DNA was extracted and quantified then genotyped using standard techniques. The highest DNA concentration of SSC samples was collected using the tape/forearm method of collection (2.1 ng/μL). Cotton swabs moistened with ethanol yielded higher quantities of DNA than swabs moistened with salicylic acid, and it gave the highest percentage of full STR profiles (97%). This study supports the use of SSC as a noninvasive sampling technique and as a extremely useful source of DNA reference samples among certain cultures where the use of buccal swabs can be considered socially unacceptable.     

人体死后肝细胞DNA含量与死亡时间关系的研究

目的研究人体死后肝脏细胞DNA含量变化与死亡时间的关系及影响因素。方法选取46例已知死亡时间的人体肝脏,根据离体肝脏所处的环境温度分为12—19％（A组）和20—27％（B组）两组,每组23例。在死后24～72h内每隔4h穿刺取肝组织1次,制成细胞悬液,经RNA酶消化,PI染色后,用流式细胞仪测定被检测细胞中含不完整DNA的细胞数所占百分比,所得数据经Exp032V1．2软件计算N值。结果死后24～72h肝细胞N平均值,A组从10．91％增至49．72％,B组从16．22％增至69．63％。两组N平均值随死亡时间的延长均逐渐增高,与死亡时间有相关性,A组r值为：0．598,B组r值为0．77357。并且建立了不同环境温度对应的回归方程。结论在不同环境温度下,死后24～72h内人体肝脏细胞DNA降解均随死亡时间的延长和环境温度的升高而逐渐加快,相关数据可望为死亡时间推断提供一定参考依据。     

Hair analysis for cocaine: factors in laboratory contamination studies and their relevance to proficiency sample preparation and hair testing practices

Since the introduction in 2001 of a urine-based detection method for recombinant erythropoietin (rHuEPO), transfusion-doping practices have regained interest. To address this problem, an efficient antidoping test designed to obtain direct proof of allogeneic blood transfusion was developed and validated. This test, based on flow cytometry analysis of red blood cell (RBCs) phenotypes, was used to determine the absence or the presence of numerous RBCs populations in a blood sample. A such, it may constitute a direct proof of an abnormal blood population resulting from homologous transfusion. Single-blind and single-site studies were carried out to validate this method as a forensic quality standard analysis and to allow objective interpretation of real cases. The analysis of 140 blood samples containing different percentages (0-5%) of a minor RBCs population were carried on by four independent analysts. Robustness, sensitivity, specificity, precision and stability were assessed. ISO-accredited controls samples were used to demonstrate that the method was robust, stable and precise. No false positive results were observed, resulting in a 100% specificity of the method. Most samples containing a 1.5% minor RBCs population were unambiguously detected, yielding a 78.1% sensitivity. These samples mimicked blood collected from an athlete 3 months after a homologous blood transfusion event where 10% of the total RBCs present in the recipient originated in the donor. The observed false negative results could be explained by differences in antigen expression between the donor and the recipient. False negatives were more numerous with smaller minor RBCs populations. The method described here fulfils the ISO-17025 accreditation and validation requirements. The controls and the methodology are solid enough to determine with certainty whether a sample contains one or more RBCs populations. This variable is currently the best indicator for homologous blood transfusion doping.     

血细胞参数与血液流变学指标对血瘀证诊断的比较

为了比较血细胞参数与血液流变学指标对血瘀证的诊断价值，为血瘀证的诊断提供新的参考指标，对９６例血瘀证患者和６０例健康人进行１１项血细胞参数和６项血液流变学指标的测定，结果显示：４项血细胞参数和５项血液流变学指标，血瘀证组显著高于对照组（Ｐ＜０．０１）；血细胞参数和血液流变学指标对血瘀证的阳性率分别为９１．７％和８７．５％，两者无显著差异（Ｐ＞０．０５）；两者对血瘀证诊断的特异性基本一致，而敏感性血细胞参数好于血液流变学指标，表明血细胞参数可作为诊断血瘀证的新参考指标。     

针刺上巨虚穴对慢传输型便秘小鼠结肠组织Cajal间质细胞数的影响

目的 探讨针刺上巨虚穴治疗慢传输型便秘（slow transit constipation,STC）的机制。方法 将40只小鼠分为正常组、模型组、上巨虚组（针刺双侧上巨虚）、非经非穴组（针刺小鼠臀部两处非经非穴点）,每组各10只。采用复方地芬诺酯灌胃法复制小鼠STC模型。观察各组首次排便时间、12 h内排便粒数,采用免疫组织化学方法观察各组小鼠结肠Cajal间质细胞(interstitial cells of Cajal,ICC)数量。结果 与正常组比较,模型组小鼠的首次排便时间明显延长 （P<0.01）,12 h内排便粒数和结肠ICC数量明显减少（P<0.01）;与模型组比较,上巨虚组小鼠的首次排便时间明显缩短（P<0.01）,12 h内排便粒数和结肠ICC数量明显增多（P<0.01）。结论 针刺上巨虚穴对STC具有明显的改善作用,其作用机制可能与升高结肠组织ICC数量有关。     

冬虫夏草菌丝水提液对SP2/0细胞凋亡的影响

在冬虫夏草菌丝水提液诱导下,采用Hoechst33342荧光染色、MTT、琼脂糖凝胶电泳和流式细胞仪等方法,观察和分析了SP2/0细胞的凋亡情况。结果显示,冬虫夏草菌丝水提液可抑制SP2/0细胞增殖,在荧光显微镜下,凋亡细胞呈亮绿色,DNA琼脂糖电泳可见典型的“阶梯状”条带,Annexin-V-FITC和碘化丙啶(PI)双染后流式细胞仪分析显示,细胞凋亡数量明显增多。表明,冬虫夏草菌丝水提液对SP2/0细胞增殖的抑制作用可能与诱导其凋亡有关。     

成年牦牛感染扩展莫尼茨绦虫的局部免疫细胞学反应

为明确扩展莫尼茨绦虫感染成年牦牛所引起的免疫细胞学反应及致病作用,对感染牛的寄生部位进行了观察与研究,并同正常成年牦牛进行了比较.结果表明,扩展莫尼茨绦虫成虫仅寄生于牦牛的小肠.寄生部位的黏膜可见少量出血点,表面附有较多黏液,散在直径2～3 mm的溃疡灶,小肠黏膜集合淋巴小结较对照组发达,数量明显增多.光镜下可见小肠黏膜出血,部分黏膜上皮坏死、脱落,局部黏膜相关淋巴组织增生明显,黏膜层嗜酸性粒细胞大量浸润,浆细胞、杯状细胞和嗜银细胞显著增生,黏膜下层血管周围肥大细胞增多.非寄生部位的组织及器官无明显病变.扫描电镜结果显示寄生部位的肠绒毛断裂或脱落,圆顶区的数量较对照组多,圆顶区滤泡相关上皮中M细胞的数量也较对照组多.研究证实,扩展莫尼茨绦虫感染可引起成年牦牛感染部位强烈的免疫细胞学反应,表明成年牦牛肠道黏膜的抗感染能力较强,故寄生虫对其致病作用较弱.     

DNA transfer during social interactions

Multiple DNA transfer has increasingly been brought up in court as potential means for the presence of the defendants DNA at the crime scene or on a piece of evidence. This has prompted several investigations into DNA transfer under very controlled and semi-controlled conditions, however little is published about DNA transfer in “uncontrolled” or real life situations.Here we examined the effects of multiple direct and indirect transfer of DNA within a small group of people and objects: three individuals participating in a social interaction of having a drink (jug of juice) together for 20 min. At the end of the tests all the surfaces of interest were sampled and analyzed.In many instances the last person or the only person to come in contact with the object was the main or the only depositor of the DNA detected on it. The jug was a clear vector for secondary DNA transfer. Interestingly, in many instances the participants acted as vectors for foreign DNA transfer.     

衣物载体上人体脱落细胞的负压吸附采集法

目的建立脱落细胞负压吸附方法,用于DNA检验中衣物等载体上人体脱落细胞的采集。方法检材包括由志愿者佩带1、5、10、20m in纱线手套,穿着5、10、20m in的内衣以及外套、鞋、帽子、头套、袜子、水杯、矿泉水瓶等。在吸尘器的进风口处连接特制的吸筒,一端覆盖具有拦截和静电吸附细胞能力的特制吸附膜,选择适当负压对各检材相应部位进行吸扫。收集吸附膜上的脱落细胞,采用Chelex-100法提取DNA,AmpFLSTR Identifiler复合扩增试剂盒扩增检测。结果上述检材用本文负压吸附法采集人体脱落细胞,经检验均获得清晰、完整的STR分型图谱,并与检材提供者的基因型一致。结论本文建立的脱落细胞负压吸附技术可有效吸附载体上的脱落细胞,适用于DNA检案实践。     

猪圆环病毒2型ORF2与EGFP融合蛋白表达载体的构建及在鸭心肌细胞中的表达

为研究猪圆环病毒2型(PCV2)ORF2基因在异源细胞上的表达及生物学特性,构建了PCV2ORF2与增强型绿色荧光蛋白(EGFP)的融合基因真核表达载体pEGFP-N1-ORF2。采用脂质体法转染体外培养的鸭心肌细胞后,通过直接荧光观察EGFP-ORF2融合蛋白在鸭心肌细胞中的分布定位,经RT-PCR、间接免疫荧光方法检测ORF2基因在鸭心肌细胞中的表达。结果显示,转染48 h后有绿色荧光出现,RT-PCR和间接免疫荧光法检测均为阳性。证实,PCV2 ORF2基因在鸭心肌细胞内获得了成功表达。