Trizol法大鼠心肌总RNA提取方法探讨

目的 探讨大鼠心肌总RNA的提取方法,为今后法医学研究和实践工作提供参考.方法 用Trizol法,严格注意无RNA酶操作,通过裂解组织,分离、沉淀RNA而提取大鼠心肌总RNA,并用分光光度计、凝胶电泳及RT-PCR检测结果.结果 Trizol法能获得较高纯度和产量的完整总RNA.结论 Trizol法是提取总RNA可靠有效的方法,可望在法医学领域有较广的应用.     

萃取方式对海洛因滥用者毛发中代谢物分析的影晌

目的比较液相萃取和同相萃取对毛发中海洛因毒品代谢物分析的影响。方法对海洛因吸食者毛发和空白添加标准品毛发经甲醇超声后的提取液分别进行液相萃取、同相萃取,然后进行衍生化和GC／MS—SIM检测。结果利用同相萃取法对添加6-单乙酰吗啡的毛发进行萃取和测试,6-单乙酰吗啡的回收率为32．O％,相对标准偏差（RSD）为2．4％;而液相萃取回收率为52．6％,相对标准偏差（RSD）为4．6％。结论固相萃取较之液液萃取,有更好的重复性,更少的杂质干扰和有机溶剂消耗等优势,但甲醇超声液需要挥干后才能进行同相萃取,而且6-单乙酰吗啡的水解率高。     

海量数据取证方法的探讨

目的对当前我国电子物证检验中提取到的海量数据信息的取证难题进行探讨。方法对数据文件中海量数据信息的特点及其传统取证方法进行分析研究。结果提出开发专用工具软件进行海量数据文件的提取。结论利用专用提取工具软件可以对常见数据文件中有规律性的数据内容进行读取、计算、查重和汇总,使海量涉案数据文件的检验从传统的人工方式转变为计算机的自动处理。     

超高效液相色谱串联质谱法检测血液中的林可霉素

目的建立超高效液相色谱串联质谱法(UPLC-MS/MS)测定全血中林可霉素的方法。方法样品血使用3mL水提取,涡旋离心,上清液过HLB柱,用10%甲醇溶液淋洗,甲醇洗脱,洗脱液用于UPLC-MS/MS分析。采用ACQUITY UPLC○R HSS T3色谱柱分离,采用电喷雾多反应监测模式(MRM)检测。结果以林可霉素母离子406.987(m/z)和子离子126.011及359.009(m/z)定性、定量。加标回收率在104.98%～120.74%。在S/N≥3的情况下,最低检出限为55.4pg/mL。结论本方法分析速度快,灵敏度高,准确度高,重现性好,可在法庭科学应用推广。     

The Effects of Differential Extraction Conditions on the Premature Lysis of Spermatozoa,

The purpose of this study was to determine the effect Proteinase K, sodium dodecyl sulfate (SDS), incubation times, and temperatures had on differential extraction efficiencies and the premature lysis of spermatozoa. The effect was measured using Quantifiler^® Duo and Identifiler? PCR Amplification kits, where the resultant male and female DNA concentrations and their ratios within the nonsperm‐ and sperm fractions (SFs) were determined. Comparisons between expected and observed ratios illustrate the quantity of female DNA in the SF increased when Proteinase K was absent during the initial incubation. Additionally, there is no indication of simultaneous sperm and epithelial cell lysis in the absence of DTT at Proteinase K concentrations ranging from 10 to 300 μg/mL. All other conditions exhibited minimal variation in DNA concentration. Therefore, despite the various protocols used for the differential lysis of cell mixtures encountered in casework, the method is robust and successful at most conditions.     

A Comparison of DNA Extraction Using AutoMate Express™ and EZ1 Advanced XL from Liquid Blood,Bloodstains, and Semen Stains

In this study, DNA was extracted using an AutoMate Express? and an EZ1 Advanced XL from liquid blood, fresh and aged bloodstains, and fresh and aged semen stains. Extracted DNA was quantified by real‐time PCR using the D17Z1 locus. Short tandem repeat typing was performed using an AmpF?STR^® Identifiler kit. The yields of DNA obtained by the AutoMate Express? were higher from fresh bloodstains and fresh semen stains, almost the same from aged bloodstains and aged semen stains, but slightly lower from liquid blood compared with those obtained by the EZ1 Advanced XL. The addition of dithiothreitol or the use of PrepFiler? lysis buffer improved the EZ1 Advanced XL results from fresh bloodstains, but not for liquid blood and aged bloodstains. Our results demonstrated that the PrepFiler? lysis buffer is the main contributor to the higher DNA yields of the AutoM ate Express? for fresh bloodstains.     

尿中吗啡的氮磷检测——气相色谱分析法

目的建立尿中吗啡的简便快速、灵敏可靠的GC/NPD分析方法。方法样品尿加内标烯丙吗啡,酶或酸催化水解,氯仿-异丙醇(9:1)液液提取或GDX403树脂固相提取,BSA衍生化,HP-5柱和氮磷检测器进行分析。结果 提取率62％～85％,检出限1.2～3.1ng/ml,线性范围20～2000ng/ml,回收率(97％～99％)±(6％~9％)(Mean±cv,N=5)。结论 方法适合于实际案件中尿样的检验。     

Enhanced elution of sperm from cotton swabs via enzymatic digestion for rape kit analysis

This report describes development of a method for enhanced cell elution from cotton swabs. The method exploits an enzyme mixture for digestion of the cotton to remove intact cells, and can be utilized in conjunction with or to circumvent conventional differential extraction (DE). Samples digested with Aspergillus niger cellulase yielded sperm cell recoveries (18+/-3.5%) similar to conventional DE buffer (23+/-7.8%) while providing intact epithelial cells. Storage time affected the concentration of enzyme required for optimal sperm cell recovery, with longer times requiring increased cellulase concentrations. Cellulase from A. niger yielded a twofold enhancement in sperm cell elution over buffer alone, and preliminary testing of higher activity cellulases from Trichoderma reesei and Trichoderma viride showed even greater enhancement. These results indicate that cellulose-digesting enzymes enhance the release of sperm and epithelial cells from a cotton swab over buffer alone, providing for efficient DNA analysis.     

关于Chelex 100提取DNA方法的进一步研究——延长保温时间对Chelex100提取DNA的影响

目的探讨用Chelex100法提取血斑DNA时,56℃保温时间、100℃保温时间长短对STR扩增的影响。方法分别改变提取过程中56℃和100℃保温时间,观察其扩增结果。结果56℃这一步骤的保温时间分别设为10min、30min和2h,100℃保温时间设为4min、8min、15min和30min,以上步骤所提取的DNA其扩增结果有明显的差异。结论在一定的时间范围内,每个STR基因座的峰值随着这两步骤保温时间的增加而增加。     

Distribution and metabolic pathway of flualprazolam in rat

Objective To probe into the distribution and metabolic pathway of flualprazolam plus its main metabolite (α – hydroxylflualprazolam) in rat. Methods 40 SD rats were randomly divided into 4 groups and fasted for 12 hours, with the first group being fed of flualprazolam by gavage as per 0.45 mg/kg and having their respective blood collected from tail vein at different time and each one’s urine received every day after administration. The second group was taken as the blank control to the first one. The third group was executed at 0.25h after drug administration, having their respective blood, heart, liver, lung, kidney, brain, spleen and muscle collected and extracted with rapid solvent extraction. The fourth group was the blank control against the third one. The distributing and metabolizing flualprazolam and α-hydroxyflualprazolam were detected through HPLC-MS/MS. Results Flualprazolam was found of rapid distributing and metabolizing in rats, reaching its highest blood concentration within 0.25h, having shown its sequential distribution in various organs as follows: kidney > brain > heart > liver > spleen > muscle > muscle > lung > blood. Yet, α-hydroxyflualprazolam demonstrated its discrepant organ-specific distributing order: liver > kidney > heart > spleen > lung > muscle > blood > brain. Conclusion The experiment has showed the distribution and degradation of flualprazolam and its main metabolite in rats who were administered of flualprazolam by gavage, capable of providing scientific basis for flualprazolam to test in vivo and have its metabolic pathway analyzed. © 2021, Editorial Office of Forensic Science and Technology. All rights reserved.