人细小病毒B19三种基因型通用核酸检测体系的建立与评价

目的：建立能同时检测人细小病毒B19（简称B19病毒）3种基因型的通用核酸检测（ NAT）体系,并进行方法学评价。方法对B19病毒3种基因型代表毒株全序列进行比对,选取其保守区域（ NS1区）设计B19病毒的通用引物和探针。为避免假阴性结果,在体系中引入竞争性噬菌体内标。将适宜浓度的内标添加到一系列梯度稀释的参考品质粒中,建立B19病毒实时荧光定量PCR（ qPCR）标准曲线。对建立的B19病毒通用NAT体系分别进行敏感性、特异性、重复性等方法学评价。结果建立的B19病毒实时qPCR检测体系的标准曲线在1×109～1× 103拷贝（copies）／μl模板浓度范围内相关性良好。方法学评价显示,体系的最低检测下限为10拷贝／μl;与其他血源传播病毒等无交叉反应;批内和批间实验重复性良好。结论建立了B19病毒3种基因型全检的通用NAT体系,不仅可用于B19病毒感染的诊断,还可用于血液和血液制品中B19病毒的NAT筛查,对于保障输血和血液制品的病毒安全性具有重要意义。     

水痘－带状疱疹病毒快速微量中和试验的建立与初步应用

目的：建立水痘－带状疱疹病毒（ VZV）的快速微量中和试验筛查技术,用于高免原料血浆的筛选和VZV特异性免疫球蛋白（ VZIG）的制备。方法参考美国FDA批准的VZIG制品,设立原料血浆筛选滴度的下限;通过病毒滴度优化,设立快速微量中和试验筛查体系中的病毒使用滴度;建立筛选体系并引入VZIG国际标准品作为质量控制,对1∶2～1∶256倍比稀释的各滴度的单人份和混合原料血浆进行测定,同时利用抗VZV中和抗体ELISA检测试剂盒VaccZyme复核快速微量中和筛选结果,对其灵敏度进行分析。结果快速微量中和筛选试验中原料血浆的最低筛选下限为1∶16（VZV中和抗体滴度不低于0．4 U／ml）;筛选体系病毒使用滴度为1500 PFU／ml;20份单采血浆中有8人份的滴度≥1∶16,混合血浆中无符合筛选标准的样本;ELISA的复核结果与快速微量中和试验呈良好的直线关系（r＝0．89524,P＜0．0001）。结论快速微量中和试验的灵敏性、通量和可操作性等,适于VZV高免原料血浆的筛选,解决了VZIG制备中原料血浆筛选的关键技术问题。     

聚合酶链反应对单-抗HBc阳性的飞行员血清HBV-DNA的检测

用聚合酶链反应对单-抗HBc阳性的飞行员血清HBV携带情况进行了调查，首先用ELISA法对189份血清检测了HBSAg、抗HBs和抗HBc。HBsAg阳性率为4．76％（9／189），抗HBs17．99％（34／189），抗HBc24．87％（47／189），聚合酶链反应检测单-抗HBc阳性血19份，抗HBs阳性／抗HBc阳性血20份。HBV－DNA阳性率分别为31．58％（6／19）和5％（1／20），结果提示单-抗HBC阳性飞行员中存在低水平的HBV携带者；在招飞体检仅检测血清HBSAg，不能除外低水平的HBV感染者。     

PreSlAg、HBeAg和HBV—DNA的对比检测与分析

目的：通过对比检测和分析739份血清中preS1蛋白、HmAg和HBV—DNA的检测阳性率，为临床正确选用实验室指标防治乙型肝炎提供依据。方法：应用ELISA法对739份血清进行PreSlAg和HBV血清标志物检测，并与实时荧光定量PCR检测HBV—DNA结果进行比较。结果：所有739份血清中，PmSlAg阳性率为34．6％（256／739），HmAg阳性率为觚7％（227／739），HBV—DNA阳性率为58.9％（435／739）。227份HBeAg阳性血清中，PreSlAg阳性率55．9％（127／227），HBV-DNA阳性率96．0％（218／227）；512份HBeAg阴性血清中，PreSlAg阳性率25．2％（129／512），HBV—DNA阳性率为42．4％（217／512）。HBeAg、HBV—DNA和PreSl的阳性率有显著差异（P〈0．05），阳性率高低依次为HBV—DNA〉PreSl〉HBeAg；HBeAg阳性血清中PreSlAg（55．9％）阳性率显著高于HmAg阴性血清PreSlAg（25．2％）阳性率（x^2=36．5，P〈0．01）；HBV—DNA阳性血清的PreSlAg检出率58．9％（256／435）显著高于HBV—DNA阴性血清的PreSlAg检出率1＆4％（56／304）（P〈0.01）；PreSl抗原与HBV—DNA阳性符合率为（200／256）7＆1％，阴性符合率为（248／483）51．3％。598份HBeAg阳性血清（阳性率为80．9%，598／739）中HBeAg、PreSl和HBV—DNA阳性数（率）分别是224（37．5%）、253（42．3oA）、417（69．7％）。结论：PreSlAg、HBV—DNA、HBeAg的阳性率有显著性差异（P〈0．05）。作为HBV感染和复制的指标，以检测HBV—DNA最为可靠，而PmSlAg可能较HBeAg更敏感。在HBV感染的不同时期，如何理解和使用上述指标对防治乙型肝炎有重要意义。     

昆明地区HBV-DNA的基因分型研究

目的：研究昆明地区乙型肝炎病毒（HBV）的基因分型特征。方法：采用荧光定量PCR及基因型特异性引物PCR方法,对昆明地区360例HBV感染血清进行HBV．DNA定量及基因分型检测,以ELISA法检测血清乙肝标志物。结果：360例HBV感染血清中,有346例（96．11％）可以直接分型,其中196例（56．65％）为C型,91例（26．30％）为B型,37例（10．69％）为B／C混合型,21例（6．07％）为D型,A型仅1例（0．29％）;比较显示,C型基因分布显著高于B和其它基因型（P〈0．05,P〈0．01）;C型基因及B／C混合基因型的HBV。DNA定量显著高于其它基因型（P〈0．01）;另除B型基因在“大三阳”模式中比例显著较低（P〈0．05）,HBV基因型在其它各血清模式组间的分布没有显著差异（P〉0．05）;14例标本未能直接分型（3．89％）,其HBV．DNA定量平均水平均低于1．00×10‘Iu／ml,显著低于其它检样（P〈0．01）。结论：昆明地区HBV基因型的分布较为丰富,存在C型、B型、B／C混合型及D型和A型,其中以C型和B型为本区域的主要HBV基因型;当B／C混合型和C型的HBV感染时,往往伴随HBV较高水平复制,且HBV感染B基因型与HBeAg的检出呈负相关;本区域内D基因型分布介于内地低海拔区域和拉萨高原地带之间。     

胰岛素自身抗体微量平板放射免疫法的建立与应用

目的建立胰岛素自身抗体（IAA）的微量平板放射免疫（RIA）法,并探讨其临床应用价值。方法^125I-胰岛素与血清在微量平板4℃保温72h后,免疫复合物转移至已包被蛋白A的Millipore过滤平板中并洗涤,于多功能液体闪烁发光分析仪上测计数。以IAA指数≥0．06作为阳性标准,通过参加第四次国际糖尿病自身抗体检测标准化评估（DASP 2005）,评价方法的灵敏度与特异性。并检测71例1型糖尿病（T1DM）患者、551例初诊2型糖尿病（T2DM）患者以及317名健康对照者IAA水平。采用SPSS 11．5软件进行t检验、非参数检验χ^2检验和直线相关分析;一致率用Kappa值评估。结果（1）优化的检测条件为5μl血清与2×10^4计数·min^-1的^125I-胰岛素于4℃缓慢振摇保温72h。（2）该方法批内CV4．8％～8．9％,批间CV6．4％～10．5％,DASP 2005反馈结果显示该法诊断T1DM灵敏度50％（25／50）,特异性97％（97／100）。与国产IAA RIA试剂盒进行比较,结果判定总体一致率为72．9％（Kappa值0．402）,相关系数（r）为0．678（P〈0．001）;26例结果不相符的标本中25例为该法检测阳性而国产RIA试剂盒检测阴性。（3）该方法检测T1DM患者的阳性率为19．7％（16／71）,显著高于健康对照组的0．9％（3／317,χ^2=54．36,P〈0．001）,其中0～9岁T1DM组9例中IAA阳性5例。检测初诊T2DM患者IAA阳性率1．5％,与健康对照组差异无统计学意义（χ^2=0．95,P〉0．05）。结论IAA微量平板RIA法灵敏度与特异性好,可应用于临床,IAA在婴幼儿T1DM患者中阳性率较高。     

孕妇乙型肝炎血清病毒标志物和HBVDNA测定及临床意义

目的探讨孕妇乙型肝炎病毒（HBV）感染者血清病毒标志物（HBV～M）组合与HBV DNA的关系以便指导免疫干预而阻断母婴传播。方法采用ELISA检测乙肝病毒血清学标志物（HBsAg、HBsAbHBeAgHBeAbHBcAb）,实时荧光定量PCR（FQ—PCR）对268例孕妇检测感染者血清中HBV DNA含量。结果HBsAg阳性组HBVDNA阳性率为49．4％（83／168）,HBsAg阴性组HBV DNA阳性率为11．10％（11／100）,两者差异显著（P〈0．01）,但HBsAg（-）组HBV DNA阳性数占总数中的11．7％（11／94）。结论临床仅根据HBV血清标志物抗原抗体的检测来判断病人的传染性是不全面的,应重视HBsAg阴性低水平HBV复制患者,结合HBV DNA的检测结果进行综合判断,以免造成宫内HBV感染。进行预防接种阻断HBV母婴传播。     

荧光酶联免疫分析技术测定人甲状腺过氧化物酶抗体

目的以重组人甲状腺过氧化物酶（hTPO）为抗原建立定量检测hTPO抗体（hTPOAb）的板式荧光酶联免疫技术（FEIA）,用于甲状腺疾病的临床检测。方法用该法测定326名健康人和各类甲状腺疾病患者279例,包括慢性淋巴细胞性甲状腺炎（HT）119例,初诊Graves病（GD）116例,结节性甲状腺肿28例,亚急性甲状腺炎10例,单纯性甲状腺肿6例的血清hTPOAb水平。组间比较采用秩和检验,组间阳性率的比较用,检验。结果（1）确立该法阳性切限值为第95百分位数（％）4000U／L（即hTPOAb浓度值大于4000U／L定为阳性）。（2）批内、批间变异系数（CV）分别为4．59％～6．52％、17．37％～17．45％。（3）该法与商品试剂盒雅培微粒荧光法检测同批血清hTPOAb结果显著相关（r=0．80,P〈0．01）。（4）用该法测定各类甲状腺疾病患者hTPOAb浓度值结果以中位数表示,HT为7000U／L,阳性率85．70％;初诊GD为4000U／L,阳性率39．65％;结节性甲状腺肿为4000U／L,阳性率28．57％;亚急性甲状腺炎为4000U／L,阳性率30．00％;单纯性甲状腺肿为3500U／L,阳性率0。HT患者血清hTPOAb阳性率与其他各甲状腺疾病组之间的差异有统计学意义（,=53．45,39．30,15．41和21．74,P均〈0．01）。结论该定量检测hTPOAb的FEIA应用于临床甲状腺疾病患者的血清学检查,在HT中有很高的阳性率,故可作为HT的诊断手段。     

不同时段介入治疗对心肌梗死室壁瘤患者心室收缩同步性及血浆脑钠肽的影响

目的对比经皮冠状动脉介入（PCI）治疗对急性心肌梗死（AMI）后左心室室壁瘤（LVA）形成、心室收缩同步性及血浆脑钠肽（BNP）的影响。方法选择2001年1月至2004年7月收治的首次急性前壁心肌梗死及左心室造影（LVG）确定合并室壁瘤者共326例,根据PCI施行的时间分为4组：A组32例（〈3h）、B组89例（≥3h且〈6h）、C组129例（≥6h且〈12h）、D组76例（AMI后1周）,4组患者于PCI后1周时行平衡法核素心室显像（ERNA）,测定左室整体和局部收缩功能、舒张功能和收缩同步性功能参数及反常室壁容积指数（PVI）;AMI后6个月随访时重复测定上述参数,并随访3年,记录主要恶性心脏事件（MACE）的发生率。所有患者于发病后18h,第5天及24周测定血浆BNP质量浓度。对数据行方差分析和,检验。结果AMI后6个月随访时,A、B、C3组左心室射血分数（LVEF）较D组明显增高（F=5．81,P〈0．05）,而相角程（PS）、半高宽（FWHM）明显降低（F=5．90和6．80,P均〈0．05）;A组反常容积消失病例数明显高于B、C、D组,且A组PVI明显低于B、C、D组[分别为（12．08±2．07）％、（15．43±2．39）％、（16．49±2．47）％、（20．41±3．68）％,F=4．32,P〈0．05]。D组发病后18h、第5天和第24周血浆BNP质量浓度均明显高于A组[（12．30±2．24）彬L与（9．85±2．60）μg／L,（9．47±1．95）μg／L与（6．65±1．56）μg／L,（5．36±1．43）μg／L与（3．27±1．12）μg／L,F=5．19,P〈0．05],B、C组差异无统计学意义（F=5．19,P〈0．05）,但均低于D组。住院期间及术后3年随访A,B,C3组梗死后心绞痛发生率和3年随访时死亡率[6．25％（2／32）与3．12％（1／32）,8．99％（8／89）与5．62％（5／89）,9．30％（12／129）与7．76％（10／129]均低于D组[21．05％（16／76）与17．11％（13／76）]∥分别为91．3和10．05,P均〈0．05。结论对AMI患者梗死相关动脉开通越早、越充分,才能越有效地抑制并逆转LVA的形成,提高左心室功能,最终改善患者预后。     

以腹水待查入院的结核性腹膜炎116例诊断体会

目的分析结核性腹膜炎（tuberculosis peritonitis,TBP）的临床特点和实验室检查．探讨各种诊断方法在TBP诊断中的应用。方法回顾分析116例以腹水待查人院并得到确诊的TBP病例,对其临床特点、实验室检查及诊断方法进行总结分析。结果各项检查的阳性率分别为PPD53．5％（38／71）,TB－Ab15．4％（2／13）。CA12592．4％（61／66）,CEA29．1％（23／79）,CA1991．7％（1／60）,ADA83．3％（20／24）,腹膜活检47．6％（10／21）,腹腔镜80％（4／5）,剖腹探查100％（10／10）。各种诊断方法诊断的例数所占比例分别为诊断性治疗77．6％（90／116）,淋巴结活检1．7％（2／116）,腹膜活检8．6％（10／116）,腹腔镜3．4％（4／116）,剖腹探查8．6％（10／116）。结论诊断性治疗是TBP确诊的主要方法。腹膜活检、腹腔镜及剖腹探查对TBP诊断的阳性率高,在有适应证的前提下应适时、及早进行。     

Knee injury and Osteoarthritis Outcome Score (KOOS) - validation of a Swedish version

The Knee injury and Osteoarthritis Outcome Score (KOOS) is a self-administered instrument measuring outcome after knee injury at impairment, disability, and handicap level in five subscales. Reliability, validity, and responsiveness of a Swedish version was assessed in 142 patients who underwent arthroscopy because of injury to the menisci, anterior cruciate ligament, or cartilage of the knee. The clinimetric properties were found to be good and comparable to the American version of the KOOS. Comparison to the Short Form-36 and the Lysholm knee scoring scale revealed expected correlations and construct validity. Item by item, symptoms and functional limitations were compared between diagnostic groups. High responsiveness was found three months after arthroscopic partial meniscectomy for all subscales but Activities of Daily Living.     

The acutely limping child

Acute limping may be the result of multiple pathologies in children. The differential diagnosis varies based on the age of the child. Irrespective of age, the initial imaging work-up includes AP and frog leg radiographs of the pelvis and ultrasound; MRI may sometimes be helpful. In children less than 3 years, infections and trauma are most frequent. MRI is the imaging modality of choice when osteomyelitis is clinically suspected. Between the ages of 3 and 10 years, transient synovitis of the hip and Legg-Calvé-Perthes disease are main considerations but infection, inflammation and focal bony lesions are also considered. In children over 10 years, slipped capital femoral epiphysis also is considered.     

Do Balance Board Training Programs Reduce the Risk of Ankle Sprains in Athletes?

Introduction Ankle sprains are the most common musculo-skeletal injury that occurs in athletes,particularly in sports that require jumping and landing on one foot such as soccer,and basketball(1-4).These injuries often result in significant time loss from participation,long-term disability,and have a major impact on health care costs and resources(5-8).     

High Intensity Interval Training:New Insights

KEY POINTS ·High-intensity interval training(HIT)is characterized by repeated sessions of relatively brief，intermittent exercise.often performed with an“a11 out”effort or at an intensity close to that which elicits peak oxygen uptake(i.e.，≥90%of VO2 peak).     

Endovascular management of carotid-cavernous fistulas

Objective To investigate endovascular treatment of traumatic direct carotid-cavernous fistulas （CCF） and their complications such as pseudoaneurysms. Methods： Over a five-year period, 22 patients with traumatic direct CCFs were treated endovascularly in our institution. Thirteen patients were treated once with the result of CCF occluded, 8 twice and 1 three times. Treatment modalities included balloon occlusion of the CCF, sacrifice of the ipsilateral internal carotid artery with detachable balloon, coll embolization of the cavernous sinus and secondary pseudoaneurysms, and covered-stem management of the pseudoaneurysms. Results All the direct CCFs were successfully managed endovascularly. Four patients developed a pseudoaneurysm after the occlusion of the CCF with an incidence of pseudoaneurysm formation of 18.2% （4/22）. A total number of 8 patients experienced permanent occlusion of the ICA with a rate of ICA occlusion reaching 36.4% （8/22）. Followed up through telephone consultation from 6 months to 5 years, all did well with no recurrence of CCF symptoms and signs. Conclusion Traumatic direct CCFs can be successfully managed with endovascular means. The pseudoaneurysms secondary to the occlusion of the CCFs can be occluded with stent-assisted coiling and implantation of covered stents.     

Developmental validation of the PowerPlex(®) ESI 16 and PowerPlex(®) ESI 17 Systems: STR multiplexes for the new European standard

In response to the ENFSI and EDNAP groups’ call for new STR multiplexes for Europe, Promega^® developed a suite of four new DNA profiling kits. This paper describes the developmental validation study performed on the PowerPlex^® ESI 16 (European Standard Investigator 16) and the PowerPlex^® ESI 17 Systems. The PowerPlex^® ESI 16 System combines the 11 loci compatible with the UK National DNA Database^®, contained within the AmpFlSTR^® SGM Plus^® PCR Amplification Kit, with five additional loci: D2S441, D10S1248, D22S1045, D1S1656 and D12S391. The multiplex was designed to reduce the amplicon size of the loci found in the AmpFlSTR^® SGM Plus^® kit. This design facilitates increased robustness and amplification success for the loci used in the national DNA databases created in many countries, when analyzing degraded DNA samples. The PowerPlex^® ESI 17 System amplifies the same loci as the PowerPlex^® ESI 16 System, but with the addition of a primer pair for the SE33 locus. Tests were designed to address the developmental validation guidelines issued by the Scientific Working Group on DNA Analysis Methods (SWGDAM), and those of the DNA Advisory Board (DAB). Samples processed include DNA mixtures, PCR reactions spiked with inhibitors, a sensitivity series, and 306 United Kingdom donor samples to determine concordance with data generated with the AmpFlSTR^® SGM Plus^® kit. Allele frequencies from 242 white Caucasian samples collected in the United Kingdom are also presented. The PowerPlex^® ESI 16 and ESI 17 Systems are robust and sensitive tools, suitable for the analysis of forensic DNA samples. Full profiles were routinely observed with 62.5 pg of a fully heterozygous single source DNA template. This high level of sensitivity was found to impact on mixture analyses, where 54–86% of unique minor contributor alleles were routinely observed in a 1:19 mixture ratio. Improved sensitivity combined with the robustness afforded by smaller amplicons has substantially improved the quantity of data obtained from degraded samples, and the improved chemistry confers exceptional tolerance to high levels of laboratory prepared inhibitors.     

Prevalence and severity of hip and groin pain in sub‐elite male football: a cross‐sectional cohort study of 695 players

The purpose of this study was twofold: (a) to investigate the prevalence of hip and groin pain in sub‐elite male adult football in Denmark and (b) to explore the association between prevalence and duration of hip and groin pain in the previous season with the Copenhagen Hip and Groin Outcome Score (HAGOS) in the beginning of the new season. In total 695 respondents from 40 teams (Division 1–4) were included. Players completed in the beginning of the new season (July–Sept 2011) a self‐reported paper questionnaire on hip and/or groin pain during the previous season and HAGOS. In total 49% (95% CI: 45–52%) reported hip and/or groin pain during the previous season. Of these, 31% (95% CI: 26–36%) reported pain for >6 weeks. Players with the longest duration of pain during the previous season had the lowest HAGOS scores, when assessed at the beginning of the new season, P < 0.001. This study documents that half of sub‐elite male adult football players report pain in the hip and/or groin during a football season. The football players with the longest duration of pain in previous season displayed the lowest HAGOS scores in the beginning of the new season.     

Bronchial stump fistula :treatment with covered retrievable hinged metallic stents-preliminary clinical experience

Objective To evaluate the preliminaily clinical efficacy and retrievability of a retrievable hinged covered metallic stent in the treatment of the bronchial stump fistula （BSF）. Methods Between April 2003 and March 2005, 8 patients with bronchial stump fistula after pneumonectomy or lobectomy were treated with two types （A and B） of retrievable hinged covered metallic stents. Type A stent was placed in 6 patients and type B in 2 under fluoroscopic guidance. The stent was removed with a retrieval set when BSF was healed or complications occurred. Results Stent placement in the bronchial tree was technically successful in all patients, without procedure-related complications. Immediate closure of the BSF was achieved in all patients after the procedure. Stents were removed from all patients but one. Removal of the stents was difficult in two patients due to tissue hyperplasia. Patients were followed up for 6 - 21 months. Placement of the stents remained stable in all patients except one due to severe cough. Permanent closure of BSF was achieved in 7 （87.5%） of 8 patients. Conclusion Use of a retrievable hinged covered expandable metallic stent is a simple, safe, and effective procedure for closure of the BSF. Retrieval of the stent seems to be feasible. （J Intervent Radiol, 2007, 16： 253-257）