鸡传染性支气管炎病毒纳米PCR检测方法的建立及初步应用     

张文  赵长润  陈基明  朱丹  张愉  王露  范文胜  磨美兰  韦平 《中国家禽》2019,(15):18-22

根据鸡传染性支气管炎病毒(IBV)的3′端非编码区保守区域,设计并合成一对特异性引物,建立IBV纳米PCR检测方法,并进行了特异性、灵敏性和重复性试验,然后将建立的方法进行初步应用。结果显示:该方法特异性良好,能从4种不同基因型IBV核酸样品中检测到约346bp的目的条带,而对禽偏肺病毒、H9亚型禽流感病毒、新城疫病毒、传染性喉气管炎病毒、传染性法氏囊病毒、马立克氏病病毒、禽白血病病毒、鸭肝炎病毒等其他病原检测结果均为阴性;敏感性试验表明,纳米PCR的最低检测浓度为1.01×10^-4ng/μL,其敏感性是普通PCR的10倍;重复性试验显示3次均能检出IBV核酸。纳米PCR方法的临床应用表明,送检样品及攻毒鸡样品的检出率分别达到50%(10/20)和60%(30/50),与普通PCR方法检测结果符合率为100%。研究建立的IBV纳米PCR检测方法具有较高的敏感性和良好的特异性,为IBV的临床诊断、病原检测和流行病学调查提供了新的技术手段。  相似文献   

Utility of an immunocytochemical assay using aqueous humor in the diagnosis of feline infectious peritonitis          下载免费PDF全文

Sandra Felten  Kaspar Matiasek  Stefanie Gruendl  Laura Sangl  Katrin Hartmann 《Veterinary ophthalmology》2018,21(1):27-34

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Comparison of histologic margin status in low‐grade cutaneous and subcutaneous canine mast cell tumours examined by radial and tangential sections          下载免费PDF全文

C.B. Dores  M. Milovancev  D.S. Russell 《Veterinary and comparative oncology》2018,16(1):125-130

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小反刍兽疫病毒核衣壳蛋白抗原性与特异性的比较     

孙雨  宋晓晖  肖颖  李秀梅  王睿男  吕园园  蒋菲  辛盛鹏  杨林  王传彬 《畜牧与兽医》2020,(5):59-63

为筛选建立小反刍兽疫病毒N蛋白双抗原夹心ELISA抗体检测方法的抗原原料,本研究通过优化大肠杆菌密码子、优化蛋白表达与纯化条件等方法,分别在pET-30a与pET-32a两个不同原核表达载体中成功获得了可溶性的小反刍兽疫核衣壳蛋白(N)。分别对重组大肠埃希氏菌pET-30a-(N)-BL21(DE3)株与pET-32a-(N)-BL21(DE3)株种子批的P7代与P20代进行蛋白诱导表达与抗原提取纯化,共获得6批小反刍兽疫病毒pET-30a-(N)蛋白与pET-32a-(N)蛋白,并对蛋白的浓度、纯度、抗原性和特异性进行检验。结果表明两个纯化后的重组抗原与羊小反刍兽疫病毒阳性血清有明显反应,具有较好的反应性,而与羊痘病毒阳性血清、羊口蹄疫病毒阳性血清、山羊关节炎/脑炎病毒阳性血清等特异性血清均无交叉反应,具有较好的特异性。本研究结果为今后以N蛋白为基础建立相应的抗原捕获ELISA方法,竞争ELISA检测方法、间接ELISA检测方法以及双抗原夹心ELISA抗体检测方法打下了坚实的基础。  相似文献   

Establishment and Application of Quantitative Real-time PCR Method to Detect Streptococcus suis Serotype 2     

LI Jian-da  YU Jiang  ZHANG Yu-yu  REN Su-fang  CHEN Lei  GUO Li-hui  SUN Wen-bo  CHEN Zhi  WANG Song  LIU Jun-zhen  DU Yi-jun  LI Jun  YANG Ling-zhi  WANG Jin-bao  WU Jia-qiang 《中国畜牧兽医》2016,43(12):3107-3113

In this study,a quantitative Real-time PCR method using the specific primers according to CPS2J gene was established to detect Streptococcus suis serotype 2. The result showed that the equation of standard curve was y=-3.073x+36.87,r=0.995,which demonstrated that the assay had good linear relationship.The melting curve analysis showed that there was only specific peak.Sensitivity test showed that the method could detect the template at the lowest concentration of 1.0×10¹ copies/μL,which was 10 times higher than the ordinary PCR.The specific tests showed that this method could able to detect Streptococcus suis serotype 2 specially and had no cross-reaction with other serotypes or other bacteria from swine. The CV of repeatability test was 0.37% to 0.63%,lower than 2.5%. The clinical diagnosis showed this assay was more sensitive than ordinary PCR and bacteria isolation. All the results showed that the established method was sensitive,specific and reproducible,which could be used for the rapid diagnosis and quantitative detection of Streptococcus suis serotype 2.  相似文献   

Receiver-operating characteristic curves and likelihood ratios: improvements over traditional methods for the evaluation and application of veterinary clinical pathology tests     

Gardner IA  Greiner M 《Veterinary clinical pathology / American Society for Veterinary Clinical Pathology》2006,35(1):8-17

Receiver-operating characteristic (ROC) curves provide a cutoff-independent method for the evaluation of continuous or ordinal tests used in clinical pathology laboratories. The area under the curve is a useful overall measure of test accuracy and can be used to compare different tests (or different equipment) used by the same tester, as well as the accuracy of different diagnosticians that use the same test material. To date, ROC analysis has not been widely used in veterinary clinical pathology studies, although it should be considered a useful complement to estimates of sensitivity and specificity in test evaluation studies. In addition, calculation of likelihood ratios can potentially improve the clinical utility of such studies because likelihood ratios provide an indication of how the post-test probability changes as a function of the magnitude of the test results. For ordinal test results, likelihood ratios can be calculated on a category-specific basis from the empirical data or by using the slope of the line joining adjacent category limits on the ROC curve. For continuous test results, data need to be categorized into intervals for estimation of likelihood ratios, or they can be calculated as the slope (tangent) to the ROC curve at a unique test value. We use ROC analysis and calculate likelihood ratios to evaluate the performance of tests reported in 2 articles previously published in this journal.  相似文献   

岩原鲤三种同工酶的组织特异性研究     

张娟  王红叶  王玉凤  蔡焰值 《湖北农业科学》2012,51(11):2307-2310

采用聚丙烯酰胺凝胶垂直板电泳,对岩原鲤(Procypris rabaudi)6种组织(眼、脑、心、肝、肾、肌肉)的乳酸脱氢酶(LDH)、苹果酸脱氢酶(MDH)、酯酶(EST)3种同工酶进行了初步研究,并对其位点及酶谱表型进行了分析.结果表明,岩原鲤3种同工酶系统具有不同程度的组织特异性.LDH检测到由3个基因位点编码,Ldh-C位点仅在肝脏组织中表达:s-MDH在6种组织中均有表达,其中在肝脏和肾脏中的活性表达较强,m-MDH在脑、心脏、肾脏和肌肉4种组织中均有表达;EST检测到由5个基因位点编码,其同工酶酶谱复杂,Est-2为6种组织的共有谱带,在肝脏中的活性表达较强.  相似文献   

EFFECTS OF OBSERVER ON THE DIAGNOSTIC ACCURACY OF LOW‐FIELD MRI FOR DETECTING CANINE MENISCAL TEARS     

Peter Böttcher  Eberhard Ludewig 《Veterinary radiology & ultrasound》2012,53(6):628-635

Low‐field MRI (lfMRI) has become increasingly accepted as a method for diagnosing canine meniscal tears in clinical practice. However, observer effects on diagnostic accuracy have not been previously reported. In this study, 50 consecutive stifle joints with clinical and radiologic evidence of cranial cruciate ligament insufficiency were investigated by lfMRI and arthroscopy. Fifteen observers who had varying levels of experience and who were unaware of arthroscopic findings independently reviewed lfMRI studies and recorded whether lateral and medial meniscal tears were present. Diagnostic accuracy (sensitivity, specificity, positive (PPV) and negative predictive value (NPV)) was determined for each observer and median values were calculated for all observers, using arthroscopy as the reference standard. Interrater agreement was determined based on intraclass correlation coefficient (ICC) analysis. Observer level of experience was compared with diagnostic sensitivity and specificity using correlation analysis. Based on pooled data for all observers, median sensitivity, specificity, PPV, and NPV for lfMRI diagnosis of lateral meniscal tears were 0.00, 0.94, 0.05, and 0.94, respectively. Median sensitivity, specificity, PPV, and NPV for medial meniscal tears were 0.74, 0.89, 0.83, and 0.79, respectively. Interrater agreement for all menisci was fair (0.51). Menisci were less consistently scored as having no tears (ICC = 0.13) than those scored as having tears (ICC = 0.50). No significant correlations between observer experience and diagnostic sensitivity/specificity were identified. Findings indicated that the accuracy of lfMRI for diagnosing canine meniscal tears was poor to fair and observer‐dependent. Future studies are needed to develop standardized and widely accepted lfMRI criteria for diagnosing meniscal tears.  相似文献   

蓝舌病病毒重组VP7蛋白单克隆抗体制备及竞争ELISA检测方法的建立     

耿宏伟  秦永丽  李俊平  杨涛  孙恩成  刘霓红  白安斌  徐青元  王凌凤  赵晶  覃绍敏  林俊  郭怡璠  吴东来 《中国预防兽医学报》2012,(5):388-392

为了建立蓝舌病(BT)的血清学诊断方法,本研究利用原核表达的蓝舌病病毒(BTV)血清型12型VP7纯化蛋白免疫BALB/c小鼠,制备2株单克隆抗体(MAb),分别命名为BTV-2D10和BTV-4H7。IFA试验表明,2株MAb均能与BTV 24个血清型发生特异性反应,而与茨城病病毒(IBAV)、中山病病毒(CV)、赤羽病病毒(AKAV)、牛病毒性腹泻病毒(BVDV)、牛传染性鼻气管炎病毒(IBRV)、牛轮状病毒(BRV)、牛肠道病毒(BEV)、牛呼肠孤病毒(RV)及口蹄疫病毒(FMDV)无交叉反应,表明2株MAb均为BTV群特异性抗体。采用重组表达的VP7蛋白作为包被抗原建立的竞争ELISA方法证明,BTV-4H7 MAb对不同血清型BTV阳性血清具有良好的阻断效果,而对AKAV、IBAV、BRV和FMDV阳性血清无阻断作用。本研究建立的竞争ELISA方法与IDEXX公司的试剂盒检测包括65份已知背景血清和322份采自广西省的山羊血清样品,检测结果符合率分别达100%和98%。该竞争ELISA方法的建立为BTV抗体的监测提供了安全、快速、准确的技术手段。  相似文献   

旋毛虫新生幼虫期特异性基因T668主要抗原表位短肽的筛选     

苗雅娟  冯爽  王学林  冯小静  于申业  唐斌  吴秀萍  刘明远 《动物医学进展》2012,(8):5-10

为进一步从T668基因编码的蛋白中寻找到敏感性及特异性较高的抗原表位,对T668主要抗原表位区进行抗原表位作图,进一步设计了一套共16条覆盖整个抗原表位决定区的短肽,并与GST融合表达,用Western blot筛选具有高反应原性的短肽。结果显示,N5EM4、N5EM5、N5EM7、N5EM8、N5EM9、N5EM116条融合短肽与感染旋毛虫的猪血清有强阳性反应条带,但均表现出较明显的假阳性。将筛选出的6条短肽人工合成后与牛血清白蛋白偶联,利用Dot blot进一步验证,结果显示融合短肽N5EM11-BSA表现出良好的反应原性和特异性。此抗原短肽的发现对进一步确定T668蛋白虫体定位、功能及旋毛虫病诊断试剂盒的研发奠定了基础。  相似文献