几种试验条件对鸡新城疫血凝和血凝抑制试验的影响

血凝抑制试验作为测定HI抗体水平,监测鸡群免疫状态,检查疫苗使用效果和制定免疫程序的合理依据已有许多报道。近年来,一些专家认为:磷酸盐对HI和HI试验有干扰作用,主张用生理盐水作稀释液。为此,本实验的目的在于观察不同pH值的生理盐水、磷酸盐缓冲液(PBS)和温度对HA和HI试验的影响,以及在同一条件下,对常量法和微量法符合情况的探讨。     

不同稀释液对禽流感H_5血清抗体检测结果的影响

本试验通过血液凝集抑制试验(HA)和血液凝集试验(HI)试验方法检测禽流感H5抗体,分别使用PBS液(0.01mol/L,p H7.2)和生理盐水作为稀释液,检测待测3份未知血清样品,1份已知阳性血清和1份已知阴性血清,每份样品做3个重复。经χ2分析发现,PBS液组和生理盐水组检测结果差异性不显著(P0.05)。试验结果证明,在日常监测中可以使用生理盐水替代PBS液作为稀释液通过血凝及血凝抑制试验检测禽流感H5血清抗体。     

影响口蹄疫正向间接血凝试验因素的研究

对影响口蹄疫正向间接血凝试验的血凝反应板，温度，稀释液的pH值及稀释液中兔血清的加入量进行了反复试验，结果表明血凝反应板只能用V型96孔110°血凝反应板，反应温度以20～25℃为宜，稀释液的pH值可控制在7．4～7．8，兔血清加入量以1％～2％为好。     

PBS液和阿氏液替代品在高致病性禽流感抗体监测中的应用初探

采用0.9%的氯化钠注射液替代PBS液和用1∶9的枸橼酸钠真空贮血管替代阿氏液进行高致病性禽流感抗体监测的血凝(HA)和血凝抑制(HI)试验。结果发现,试验结果均有效。且采用这种替代品稳定、方便、省时、省工。     

影响血凝抑制试验的因素

1稀释液 最常用的稀释液为生理盐水,要求其无菌、无杂质.试验前要先调整其pH为7.0.若稀释液生理偏酸性,易使红细胞溶解;偏碱时,吸附于红细胞上的病毒易脱落.     

榆中地区鸡禽流感H5亚型的血清学调查

2007年4月底,笔者在榆中县进行鸡禽流感春检工作,对23个乡镇的344份血清,应用血凝抑制试验(HI)进行了血清学检测。结果显示,应用于本次血凝抑制(HI)试验的4个血凝单位抗原的稀释倍数为512倍(29);在检测的阳性结果中,鸡禽流感H5亚型HI最高抗体滴度为512(29),主要分布在青城﹑甘草店﹑园子3个乡镇;HI最低抗体滴度为32(25),主要分布在中连川﹑和平﹑上花3个乡镇。血清抗体阳性率以青城乡最高,达100%,血清抗体阳性率以来紫堡乡最低,为35.71%;榆中县总体血清抗体阳性率为74.42%。由此可见,H5亚型禽流感病毒阳性率较高,且抗体滴度也较高,说明该地区大多数家禽对禽流感H5亚型病毒可以产生较强的保护抗体。     

影响血凝及血凝抑制试验结果因素的分析

血凝和血凝抑制试验是兽医实验室中进行动物疫病诊断和免疫监测最常用的试验方法之一,经常用到如鸡新城疫、禽流感、口蹄疫、猪瘟等多种疫病的检测试验中。石家庄市各级兽医实验室每年春防、秋防后对鸡新城疫、禽流感等主要动物疫病均进行免疫抗体集中监测,以便准确掌握免疫质量。因此血凝和血凝抑制试验对鸡新城疫等主要动物疫病的监测就显得尤为重要。但血凝和血凝抑制试     

苏州地区高致病性禽流感抗体水平监测

禽流感，尤其高致病性禽流感对养禽业的危害十分严重。禽类在紧急免疫后的抗体水平直接关系疫情的控制、养禽业的发展以及人类的健康。因此我们用血凝及血凝抑制试验对高致病性禽流感抗体水平进行监测显得十分重要和必要。     

浅析影响血凝和血凝抑制试验的因素

血凝和血凝抑制试验是化验室中进行动物疫病诊断和免疫监测最常用的试验方法之一，如禽流感、口蹄疫等疫病的化验室诊断或免疫监测在基层多使用这种试验方法。今年在我国其它省市发生多起禽流感等重大动物疫病威胁我省我市的情况下，省、市、县等各级畜牧兽医部门都对禽流感进行了大量的化验室监测，运用和准确掌握血凝和血凝抑制试验对禽流感等重大动物疫病的诊断和免疫监测就显得尤为重要。但血凝和血凝抑制试验方法在操作中常有较多的影响因素，容易造成试验结果偏差，甚至误判。现将这种实验方法操作中影响因素浅析如下。     

影响禽流感血凝和血凝抑制试验的因素及对策

血凝和血凝抑制试验是目前较为广泛使用的监测禽流感抗体水平的一种试验方法.对血凝和血凝抑制试验中的影响因素及对策进行了阐述.     

Hemagglutination and hemagglutination inhibition with Chuzan virus.

Chuzan virus agglutinated erythrocytes of several species of animals including bovine. The hemagglutinating (HA) activity against bovine erythrocytes was dependent on NaCl molarity and was expressed best at 0.6 M, but it was independent of pH and temperature. Three strains of Chuzan virus isolated from 2 cows and a pool of culicoides midges had indistinguishable HA antigenicity. All cattle infected with the virus developed high titers of hemagglutination inhibiting (HI) antibody which changed in parallel with neutralizing (NT) antibody titers. Correlation between HI and NT antibodies was very high and the antibodies persisted for one year or more. Therefore it was concluded that the HI test is applicable for survey of Chuzan virus infection among cattle in place of the NT test.     

Evaluation of a gel diffusion precipitin test for porcine parvovirus

The use of a gel diffusion precipitin (GDP) test for the detection of porcine parvovirus (PPV) infection in pigs is described. The close correlation between gel diffusion precipitin and haemagglutination inhibiting (HI) antibody titres indicates that, with careful standardisation, a high level of sensitivity can be achieved with the GDP test and that it is a simple and relatively inexpensive alternative to the more commonly used HI test. Experimental infection of 2 groups of pigs showed that GDP and HI antibody responses were closely correlated and that GDP antibodies to PPV persisted for at least 41 weeks after infection. In a commercial herd study, serological evidence of declining passive immunity and subsequent acquisition of active immunity was demonstrated by measuring the GDP and HI antibody titres in sequential serum samples of pigs from a known PPV endemic farm. The GDP test described was shown to be less sensitive than haemagglutination (HA) in the detection of viral antigen but was, nevertheless, considered useful as a simple screening test for the amounts of antigen usually present in PPV infected mummified foetuses.     

Specificity of purified protein derivative extracts from ten species of Mycobacteria killed with phenol in the haemagglutination and haemagglutination inhibition test

The specificity of purified protein derivative (PPD) extracts from 10 species of Mycobacteria killed with phenol was evaluated using the haemagglutinin (HA) and haemagglutination inhibition (HI) tests. The HA test showed many cross-reactions with homologous and heterologous antisera. In the HI test, many antisera cross-reacted with most PPD extracts. The PPD extracts from Mycobacterium foruitum and M. smegmatis were the most specific. It was concluded that PPD extracted from unheated bacilli was not sufficiently specific for use in HI tests.     

3种犬细小病毒感染诊断方法比较

本研究应用犬细小病毒抗原快速检测试剂盒、血凝/血凝抑制试验和PCR方法对120份具有腹泻/呕吐症状的患犬粪便样本进行了犬细小病毒检测。结果显示,3种方法检测阳性份数分别为60,49,68。犬细小病毒抗原快速检测试剂盒与血凝/血凝抑制(HA/HI)试验相比较,敏感性、特异性及总符合率分别为100.0%,84.5%,90.8%;与PCR法相比较,敏感性、特异性及总符合率分别为85.3%,96.2%,90.0%。从试验结果看,HA/HI法具有较高的特异性,但敏感性较低;PCR具有高度敏感性和特异性,但不适合临床推广;细小病毒抗原快速检测试剂盒,在正确操作下结果可靠,是临床诊断犬细小病毒感染的首选方法。     

利用不同源红细胞检测鹅禽流感抗体效价的报告

从某一已免疫禽流感的种鹅场采取30份血样，分别以健康鸡红细胞悬液和健康皖西白鹅红细胞悬液为指示剂，对30份血样进行禽流感血凝和血凝抑制试验。结果：用鸡红细胞悬液作指示剂进行血凝试验，测出抗原4HAU为1：32；用鹅红细胞悬液作指示剂进行血凝试验，测出抗原4HAU为1：8以1：32倍数稀释抗原，分别用鸡红细胞和鹅红细胞悬液作指示剂进行血凝抑制试验，测出30份鹅血清禽流感H5免疫抗体效价的平均值分别为2^5.8和2^3.9；以1：8倍数稀释抗原，用鹅红细胞悬液作指示剂进行血凝抑制试验，测出30份鹅血清禽流感免疫抗体的平均效价为2^5.1。     

Protection studies on winter dysentery caused by bovine coronavirus in cattle using antigens prepared from infected cell lysates.

Cells infected with bovine coronavirus (BCV) were solubilized with Triton X-100 to yield a cell lysate (CL) antigen having high hemagglutinating (HA) titers. The antigen gave high HA titers using rat erythrocytes, suggesting that it contained large amounts of hemagglutinin esterase (HE) antigen. The CL antigen, combined with an oil adjuvant, was tested for protective and antibody-inducing activities in cattle. Four groups (2 cattle/group) of cattle were inoculated with CL antigen having HA titers of 16 000, 4000, 1000, and 250. Another group served as untreated controls. Two intramuscular inoculations were given at an interval of 3 wk. The animals were challenged with virus 1 wk after the second inoculation. The groups immunized with the CL antigen having an HA titer of 4000 or 16 000 produced hemagglutination inhibition (HI) antibody titers of > 320 and serum neutralizing (SN) antibody titers of > 1280. These groups of animals showed no clinical abnormalities after challenge. In the groups immunized with CL antigen at an HA titer of 1000 or 250, HI antibody titers were 40 to 160 and SN titers were 80 to 640. The cattle with HI antibody titers of > or = 160 and the SN titers of > or = 640 showed no clinical signs, but the cattle with the HI antibody titer < 80 and the SN antibody titer < 160 developed watery diarrhea and fever after challenge. These results indicate that CL antigen with high HA titer induces antibody production in cattle that provides effective protection against winter dysentery.     

B型副鸡嗜血杆菌血凝素基因的克隆表达及生物活性鉴定

血凝素是副鸡嗜血杆菌（Haemophilus paragallinarum，Hpg）的主要抗原成分之一，鸡只对Hpg的免疫力与血凝素抗体滴度成正相关。根据GenBank上已发表的B型Hpg Dalian株的血凝素基因序列（AY622378），设计合成了1对特异扩增Hpg血凝素基因的引物。以大连分离株Hpg中提取的细菌DNA为模板，利用PCR扩增出了1038bp的血凝素基因，将其克隆到pET-32a载体上，构建了pET-HA原核表达质粒并在BL21。大肠杆菌中过量表达。血凝试验显示纯化的重组蛋白具有血凝活性；Western试验证明该重组蛋白可以被B型Hpg特异性鸡血清所识别。本研究在国内首次对Hpg的血凝素基因进行克隆表达，并分析了重组蛋白的血凝活性。     

Preparation and diagnostic utility of a hemagglutination inhibition test antigen derived from the baculovirus-expressed hemagglutinin-neuraminidase protein gene of Newcastle disease virus

A recombinant hemagglutinin-neuraminidase (rHN) protein from Newcastle disease virus (NDV) with hemagglutination (HA) activity was expressed in Spodoptera frugiperda cells using a baculovirus expression system. The rHN protein extracted from infected cells was used as an antigen in a hemagglutination inhibition (HI) test for the detection and titration of NDV-specific antibodies present in chicken sera. The rHN antigen produced high HA titers of 2¹³ per 25 µL, which were similar to those of the NDV antigen produced using chicken eggs, and it remained stable without significant loss of the HA activity for at least 12 weeks at 4℃. The rHN-based HI assay specifically detected NDV antibodies, but not the sera of other avian pathogens, with a specificity and sensitivity of 100% and 98.0%, respectively, in known positive and negative chicken sera (n = 430). Compared with an NDV-based HI assay, the rHN-based HI assay had a relative sensitivity and specificity of 96.1% and 95.5%, respectively, when applied to field chicken sera. The HI titers of the rHN-based HI assay were highly correlated with those in an NDV-based HI assay (r = 0.927). Overall, these results indicate that rHN protein provides a useful alternative to NDV antigen in HI assays.     

Hemagglutination and antigenic comparison of rabbit hemorrhagic disease virus

The hemagglutinating activity and serological properties of three strains of rabbit hemorrhagic disease virus, Chinese, Korean and Shizuoka, which was first isolated in Japan, were examined by hemagglutination (HA) and cross hemagglutination inhibition (HI) test with human erythrocytes. Similar results were observed between the Chinese and Korean strains, both of which gave positive HA at 4 degrees C with O, A, B and AB, and at 22 degrees C with B and AB blood groups. In the Shizuoka strain, positive HA was observed at 4 degrees C with O, A, B and AB, at 22 degrees C with A, B And AB, and at 37 degrees C with B blood group. In experimentally infected rabbits, HI antibody in these animals showed a titer of 16,384 or 32,768 at 4 weeks after inoculation. No serological difference was observed in three strains by cross HI test.     

新版国家标准《新城疫诊断技术》血凝和血凝抑制试验的理解与操作实践

血凝（HA）和血凝抑制试验（HI）主要是用于新城疫病原鉴定和抗体效价测定,新版《新城疫诊断技术》（GB/T16550-2020）对HA和HI试验判定结果进行了修改和细化,使其更加标准和规范。为尽快理解新版标准要求,熟练掌握操作要领,对新、旧版标准中HA和HI试验进行了比较,新版标准增加了对HA和HI试验4HAU抗原配制的标定、静置时间以及结果判定的改变。新版标准在操作步骤上更为精准详细,4HAU抗原配置更加准确,从而使得HI试验结果更加科学。本文结合实际检测,为广大实验室检验人员、大专院校学生及科研人员更好地理解新版标准,更熟练地操作,从而为提高试验结果准确性提供了参考。