口蹄疫病毒O、A、AsiaⅠ型定型诊断胶体金免疫层析方法的建立

【目的】建立一种从田间样品中快速、灵敏、简便检测O、A、Asia I型口蹄疫病毒抗原的胶体金免疫层析方法（GICA）。【方法】采用柠檬酸三钠还原制备胶体金颗粒,标记纯化的标准株O、A、Asia I型口蹄疫抗体。将纯化的口蹄疫流行株O/China99,A/AF72 and AsiaⅠ/China05抗体和羊抗豚鼠抗体分别包被在硝酸纤维素膜上,作为检测带与质控带。经试验条件优化,组装形成胶体金定型诊断试剂盒。【结果】本试验研制的口蹄疫定型试剂盒具有良好的灵敏度,可检测到7.8×104 LD50（1﹕128倍稀释乳鼠毒）的病毒量。同时对阳性及阴性样品进行3次重复检测结果完全相同,说明其有较好的重复性;特异性试验证实该方法在检测口蹄疫临床症状相似病原,如猪水疱病抗原（SVD）、水泡性口炎（VS）、水泡性疹（VES）等病毒时无交叉反应;试剂盒对206份已知血清型田间及实验室样品的符合率试验,O、A、AsiaⅠ型和阴性样本的符合率分别为92.45 %、91.66%、92.75%、100%定型准确率94.58%。试剂盒在4℃下可保存6个月。【结论】本试验研发的口蹄疫病毒胶体金定型诊断试剂盒是一种快速、灵敏、特异的FMD抗原检测方法,对基层现场具有广泛应用价值。     

柞蚕微孢子虫胶体金免疫层析检测法

利用柞蚕微孢子虫孢子液直接免疫家兔获得柞蚕微孢子虫多克隆抗体后,分别采用双抗夹心法和竞争法制作胶体金免疫层析试纸条,建立能简便、快捷、准确诊断柞蚕微粒子病的柞蚕微孢子虫胶体金免疫层析检测法。双抗夹心法和竞争法分别是将柞蚕微孢子虫多克隆抗体与25 nm和17 nm的胶体金颗粒结合并固定在金标垫上,然后均将羊抗兔二抗包被在硝酸纤维素膜上作为质控点(C),但是2种方法制作的胶体金免疫层析试纸条的反应模式不同:前者是以柞蚕微孢子虫多克隆抗体作为检测点(T),而后者以柞蚕微孢子虫孢壁蛋白作为检测点(T)。2种方法制作的胶体金免疫层析试纸条的检测时间均为10 min,检测灵敏度为0.8×107个/mL,与柞蚕血淋巴无交叉反应,特异性强,操作简单,其中,以双抗夹心法制作的试纸条显色更清晰,结果更可靠,更具实用价值。     

Usefulness of a rapid immuno-migration test for the detection of canine monocytic ehrlichiosis in Africa

A rapid immuno-migration test for the serological detection of canine monocytic ehrlichiosis, Witness^® Ehrlichia (WE) (Zoetis, France), was evaluated in 528 serum samples from dogs living in endemic areas of West and East Africa: Senegal (N = 208), Ivory Coast (N = 7), Sudan (N = 27), and Djibouti (N = 286). Of these dogs, 200 were French military working dogs (MWD) temporarily residing in Africa. The WE test results were compared with those obtained by indirect immunofluorescence (IFA). The sensitivity of WE was 97% [94.2, 98.7] with a specificity of 100% [98.6, 100]. In MWD, the seroprevalence (IFA) was 7%; in native dogs, it reached 77.1%. This significant difference can be explained by prophylactic measures from which MWD benefit. The WE test represents a simple, fast and reliable test for the detection of anti-Ehrlichia canis antibodies. Its implementation for the diagnosis of clinical cases has been validated in the field, and its use allows easy detection of asymptomatic dogs that may be carriers of E. canis.     

2种检测粮食中呕吐毒素的免疫技术比较

对检测粮食中呕吐毒素的2种免疫技术,免疫亲合柱(IAC)-HPLC)法和免疫色谱法(速测卡)进行了比较,结果表明I:AC-HPLC法的最低检测限可达到0.1 mg/kg,相对标准偏差为8.76%～9.82%,回收率87.3%～93.6%,方法灵敏、准确;免疫色谱法的最低检测限为0.5 mg/kg,操作快速、简便,适合初筛。     

A Bayesian evaluation of four immunological assays for the diagnosis of clinical cryptosporidiosis in calves

A Bayesian approach was used to evaluate four immunological assays for the clinical diagnosis of cryptosporidiosis in calves: an immunofluorescence assay (IFA), two ELISA tests and an immunochromatographic (dipstick) assay. Faecal samples from 287 calves aged less than 6 weeks with clinical signs of gastrointestinal disease were examined for the presence of Cryptosporidium spp. The high prevalence (63%) of Cryptosporidium spp. indicated the relevance of this agent in the aetiology of diarrhoea in calves. All diagnostic assays were found to be relatively specific (IFA: 94.8%; Tetra ELISA: 95.9%; Techlab ELISA: 92.7%; dipstick assay: 91.5%) and sensitive (IFA: 97.4%; Tetra ELISA: 93.6%; Techlab ELISA: 95.4%; dipstick: 87.8%). Despite a lower sensitivity, the dipstick assay provided a practical alternative to laboratory diagnosis of clinical cryptosporidiosis in calves.     

快速检测联苯菊酯的胶体金层析试纸条研制

[目的]采用免疫胶体金技术,建立一种快速检测联苯菊酯的试纸条方法。[方法]利用有机合成方法制备了联苯菊酯半抗原,并与载体蛋白偶联得到免疫抗原,利用人工免疫的方法获得其单克隆抗体,通过酶联免疫法对制备的单克隆抗体的性能进行了验证。联苯菊酯胶体金层析试纸条是利用单克隆抗体标记胶体金,并将联苯菊酯抗原与羊抗鼠Ig G分别喷制到硝酸纤维膜上,与样品垫、吸水垫组装在PVC衬板上制备而成。[结果]联苯菊酯胶体金层析试纸条检测限为500~800μg/kg,具有速度快、简单等优点。此外,该试纸条与大部分的拟除虫菊酯没有明显的交叉反应,利用该试纸条可实现对苋菜中联苯菊酯添加试验的准确判定。[结论]胶体金试纸条方法具有前处理简单、检测速度快、可以肉眼判断结果等优点,该试纸条有望实现对果蔬中联苯菊酯残留的快速筛查。