Ovine campylobacterosis: preliminary studies of the efficacy of the in vitro serum bactericidal test as an assay for the potency of Campylobacter (Vibrio) fetus subsp intestinalis bacterins.

An in vitro serum bactericidal test was developed to assess the efficacy of Campylobacter fetus bacterins. Four experimental monovalent bacterins (either serotype C or A-2) and 2 commercial bivalent bacterins (a "suspect" and an "efficacious" bacterin) with aluminum hydroxide adjuvant were administered to sheep and rabbits from which antiserums were then prepared. The different vaccines were evaluated by comparing the in vitro bactericidal activity of the sheep and rabbit antiserums. Results of the in vitro tests were compared to the protection induced in vaccinated ewes which were orally exposed to C fetus. The sheep and the rabbit antiserums after they were heated at 56 C for 30 minutes were unable to exert a killing effect on C fetus cells. Addition of a fresh homologous complement source to the heated antiserums was necessary to demonstrate the in vitro bactericidal capacity. In the comparison of the suspect and the efficacious commercial bacterins, which both reportedly contain serotype C cells, there was a statistically significant difference in bactericidal activities for serotype C cells of antiserums from sheep 14 days after the 2nd vaccination. There was a corresponding significant difference in the antiserums from rabbits 14 days after the 2nd vaccination. Proportionally, more abortions and stillbirths were observed in the ewes vaccinated with the suspect bacterin and then orally exposed to C fetus-serotype C cells than in those vaccinated with the efficacious bacterin. The results indicated that the ability of vaccinated sheep to overcome infection is reflected in the in vitro bactericidal capacity of antiserum from the animal. Since 89% of the variation in sheep antiserums from 14 days after 2nd vaccination can be accounted for by rabbit antiserums from 14 days after 2nd vaccination, the in vitro bactericidal capacity of rabbit antiserums probably provides a reliable index of the protective effect of bacterins containing serotype C for ewes exposed to the homologous serotype.     

Quantitation by ELISA of pili and sheep antibodies to the pili of Bacteroides nodosus

ELISA systems have been developed to quantitate the isotypic antibody response of sheep naturally infected with B. nodosus isolate 198 or injected with pili from isolate 198 in oil emulsion vaccines. The predominant humoral antibody detected following vaccination was IgG1, with substantially lower amounts of IgG2 and IgM. The antibody response was relatively specific for the pilus antigen from isolate 198. Although weak cross reactivities were detected with antiserums to some other isolates, ELISA IgG antibody titres in excess of 200 offer a tentative identification of the isolates of B. nodosus involved in natural outbreaks of footrot. A related ELISA was also developed to quantitate the amount of pili in cell suspensions and crude preparations of pili used in vaccines.     

鸡L-FABP抗血清制备及组织表达特性分析

为制备鸡肝脏型脂肪酸结合蛋白（L-FABP）的多克隆抗血清,并分析L-FABP的组织表达特性,利用RT-PCR扩增L-FABP基因,构建鸡L-FABP基因的GST融合蛋白表达质粒pGEX-4T／L-FABP。将重组表达质粒转化大肠杆菌BL21,IPTG诱导产生GST／L—FABP融合蛋白,用亲和层析纯化目的蛋白,将纯化的GST／L—FABP融合蛋白免疫家兔制备抗血清,并利用此抗血清分析鸡0FABP基因的组织表达特性。诱导得到了1个40ku（14ku L—FABP＋26kuGST）的融合蛋白,获得效价较高、特异性强的鸡L-FABP的抗血清。鸡L-FABP的组织表达特性研究结果表明,该基因在肝脏和小肠组织中有较高表达,但在心脏、脂肪、肌肉、肌胃、脾、肺和肾中没有检测到表达信号。     

Haemophilus somnus complex: antigenicity and specificity of fractions of Haemophilus somnus.

Five Haemophilus somnus type 8025 preparations (whole cell, sonicate, crude polysaccharide, purified polysaccharide, and protein) were produced for studies of their antigenicity in rabbits. Bacterial agglutination and passive hemagglutination tests were used to assess the level of antibody produced in rabbits inoculated with the different antigenic preparations. Cross-reactions were seen between the antiserums against the H sumnus 8025 antigens and a variety of related and unrelated bovine pathogens. The strongest cross-reaction occurred between antiserums against H somnus 8025 whole cell and crude polysaccharide antigens and Haemophilus agni and Actinobacillus lignieresii cell suspensions.     

Serological characterisation of Australian isolates of Actinobacillus pleuropneumoniae

SUMMARY: Using hyperimmune rabbit antiserums to 8 reference serovar strains, a total of 51 Australian isolates of Aclinobacillus pleuropneumoniae were serotyped by either a rapid slide agglutination test or a gel diffusion test. The results were: serovar 1 — 24 isolates; serovar 2 — 6 isolates; serovar 3 — 5 isolates; serovar 7 — 15 isolates; nontypable — 1 isolate. The rapid slide agglutination test was suitable for screening field isolates, as 73% of those tested reacted with only 1 of the 8 antiserums and were assigned to a particular serovar of the basis of this test alone.     

Haemophilus somnus as a cause of bronchopneumonia in calves

A bacteriological examination of the pulmonary tissues of calves suffering from bronchopneumonia was complemented by a microaerophilic or anaerobic cultivation. Thirty-two strains (23.7%) of a microorganism hitherto not described in Czechoslovakia were isolated during the examination of 135 calves with disorders of the respiratory system. On the basis of the growth, morphological, biochemical and serological properties, the microorganism was identified as Haemophilus somnus. Chocolate agar supplemented with yeast extract, and blood agar were used for the cultivation. Incubation took place in a microaerophilic medium with 10% carbon dioxide or in an anaerobic medium. Serums against the collection strain of Haemophilus somnus were prepared in rabbits for the serological diagnosis. These antiserums reacted with all tested strains during test tube agglutination but did not enter cross reactions with other bacterial species. The coagglutination test, which was verified, did not provide reliable results.     

Serological Studies of African Horse-Sickness Virus with Emphasis on Neutralization Test in Tissue Culture

In place of mice, monkey kidney stable (MS) cell cultures were used successfully in serologic studies of African horse-sickness virus.

The maintenance medium containing 2% serum was chosen as the virus diluent. Maximum neutralization occurred after 1-hour incubation at 37 C., and maintained the same titer during an additional 4-hour incubation period. No significant difference was observed between neutralization titers titrated using the same antiserum mixed with two different passage levels of virus.

Rabbit and guinea pig antiserums prepared using virus grown in MS cell cultures had antibody titer as high as those prepared in the same manner using mouse brain suspension.

African horse-sickness virus strains isolated in Asia were serologically identified using a standard neutralization technique in tissue culture. All the strains were closely related to each other and all had antigenic similarity to Type 6 virus (strain 114).

     

Identification of various porcine mycoplasmas using epi-immunofluorescence]

The responses of the field strains and type strains of Mycoplasma hyorhinis are compared in epi-immunofluorescence, in growth-inhibition tests, and growth-precipitation tests. The type strains, together with the field strains, could be definitely identified by direct and indirect epi-immunofluorescence. In indirect epi-immunofluorescence, all rabbit antiserums against the strains of Mycoplasma hyorhinis tested by us always gave positive cross reactions with all the tested strains of Mycoplasma hyorhinis. On the other hand, in the growth-inhibition cross test and growth precipitation cross test, the type strains of Mycoplasma hyorhinis always reacted positively only with the homologous antiserums, whereas the heterologous reactions were mostly negative or dubious. The clearly positive epi-immunofluorescent reactions with serums against any strain of Mycoplasma hyorhinis suggest that this method is more suitable for the identification of Myocplasma species than the other two tests studied.     

Immune electron microscopy of transmissible gastroenteritis virus and rotavirus (reovirus-like agent) of swine.

Immune electron microscopy (IEM) was developed as a diagnostic aid for detecting and identifying transmissible gastroenteritis virus and rotavirus (reovirus-like agent) in fecal and intestinal contents from cases of gastroenteritis in young pigs. Variables involved in use of direct IEM and its sensitivity were determined. Aggregates of virus coated with specific antibody were seen in virus samples mixed with homologous convalescent antiserum, but not in control samples containing preexposure serum or antibody directed against a heterologous virus. At least a ten fold enhancement of the sensitivity of direct IEM for virus detection was accomplished using indirect IEM employing rabbit anti-porcine IgG to further aggregate virus-antibody complexes. The technique was used to investigate the size and morphology of the porcine rotavirus. Particles ranged from 55 to 70 nm in diameter and had capsomere structures. Morphologically, the porcine rotavirus resembled the calf and human rotaviruses. By IEM, employing specific antiserums for each virus, porcine rotavirus was found to be antigenically related to these 2 viruses, but not to the reovirus type 3.     

Purification of IgG from serum with caprylic acid and ammonium sulphate precipitation is not superior to ammonium sulphate precipitation alone

Immunoglobulin G (IgG) from bovine serum raised against Aeromonas Salmonicida was purified by ammonium sulphate precipitation (ASP) or caprylic acid treatment followed by ammonium sulphate precipitation (CAAS). Purity of IgG samples prepared by both methods were examined by High Performance Gel Permeation Chromatography, electrophoresis and antibody activity assay. Results suggest that IgG prepared by ASP is better than that obtained by CAAS method in terms of the yield of the IgG monomers and the recovery of the antibody activity.     

鸡免疫球蛋白G Fc(IgG Fc)片段的分离纯化及其抗血清的制备 Ⅱ.豚鼠抗鸡IgG Fc片段血清的制备

经硫酸铵盐析、DEAE 32-纤维素和Sephadex G 200柱色谱法分离得到纯化的鸡血清IgG。然后用木瓜蛋白酶水解IgG,再经DEAE 32-纤维素柱色谱纯化制得IgG Fc片段,并以IgG Fc片段免疫豚鼠制备豚鼠抗鸡IgG Fc血清。     

Immunocompetent cells of the turkey: antigenic surface determinants of turkey lymphoid cells.

The serologic properties of chicken antiserums to turkey bursa and thymus were assayed by the cytotoxicity tests and indirect immunofluorescence. The following antigenic surface determinants were detected, using proper absorptions on thymic and bursal lymphoic cells: (a) common lymphocyte antigens present on both kinds of cells, (b) thymus-specific antigens, (c) bursa-specific antigens, and (d) immunoglobulin surface determinants in bursa cells, as revealed by direct immunofluorescence.     

Immunoglobulin G1 Fc in colostral whey.

Two IgG1 fragments were isolated from bovine colostral whey. Based on gel diffusion analysis, both fragments originated from the Fc portion of the molecule but were not identical to IgG1 Fc prepared by papain cleavage. The molecular weights were determined to be 66,000 and 14,000 daltons and it was hypothesized that the larger fragment could be a polymer of the smaller. No IgG1 Fab or IgG2 fragments could be demonstrated.     

人血清中分离纯化鸡IgG、IgM的实用方法

本试验用绵羊红细胞免疫鸡,免疫后5天采血,制备富集IgM血清。应用硫酸铵粗提、Sepharose-4B分子筛层析方法,从富集IgM的鸡血清一次性分离提取高纯度的IgM及纯度较高的IgG,DEAE－52纤维素离子交换层析进一步纯化,获得电泳纯的IgG。经鉴定该方法制备的IgG,IgM完全可以满足免疫学实验要求,从一份血清样品中同时纯化IgG、IgM两种免疫球蛋白,既省时又节省原料。     

Evaluation of enzyme-linked immunosorbent assay for quantitation by subclass for bovine antibodies.

The enzyme-linked immunosorbent assay (ELISA) was evaluated for use in the quantitative measurement of bovine immunoglobulin IgG1 and IgG2 antibodies. A method for standardization was devised in which IgG1 or IgG2 was directly adsorbed to polystyrene tubes and the actual degree of binding was calculated by using different input amounts of 125I-labeled IgG1 or IgG2. Values for quantity of IgG1 antibodies to human serum albumin were only slightly higher when measured by the ELISA than when measured by quantitative precipitation although the value measured by the ELISA for IgG2 antibodies was twice that determined by quantitative precipitation. This discrepancy could result from conjugate cross reactivity, differences in affinity between antibodies of the 2 subclasses, or the occurrence of IgG2 nonprecipitating antibodies. The danger of overlooking subclass anti-globulin cross reactivity because of the failure to detect it by immunoprecipitation, also is illustrated. In addition, only enzyme-antibody conjugates prepared with specifically purified antibodies were effective, and reproducibility of individual data points required that 4 replicate determinations be performed. Advantages, pitfalls, and limitations of the ELISA are discussed.     

牛初乳乳清制备工艺研究

为优化乳清制备工艺,以脱脂牛初乳为原料,研究通过酸沉酪蛋白方法制备高质量乳清。综合考虑酸的种类、强弱和乳清中IgG的活性变化,确定了酸沉酪蛋白的最佳工艺条件为：0．8mol／L乳酸调脱脂乳pH值至4．3,加水至干物质终质量分数为10％,45℃水浴15min,4000r／min离心5min。经检测所制备的乳清中IgG活性含量仅损失3．05％。     

Application of radioimmunological methods for checking the quality of class-specific antibodies against bovine and porcine immunoglobulins

Class-specific antibodies against bovine IgG1, IgG2, IgM and IgA and porcine IgG, IgM and IgA immunoglobulins were prepared. Their class specificity was assessed by two radioimmunological methods, namely, radioimmunoelectrophoresis and double antibody sandwich radioimmunoassay. The methods are highly specific and sensitive and do not require the use of purified immunoglobulins, but can be performed with normal serum or colostrum. It was confirmed that antibodies found satisfactory in these tests were suitable for a wide range of use including radioimmunoassay and enzyme linked immunosorbent assay.     

鸡免疫球蛋白(GIgG)轻链的分离纯化及其抗血清的制备

经硫酸铵盐析、柱色谱分离得到纯化的鸡血清IgG。继用2-巯基乙醇还原、碘乙酰胶烷化拆开IgG的轻、重链,再经Sephadex G100凝胶过滤柱色谱分离得到IgG轻链。以IgG轻链免疫家兔制得兔抗鸡IgG轻链抗血清。     

Isolation and characterization of epizootic hemorrhagic disease virus from white-tailed deer (Odocoileus virginianus) in eastern Washington.

A virus was isolated from the spleen of a white-tailed deer (Odocoileus virginianus) that had died during an epizootic in Washington state in 1967. Inoculation of a 10% spleen suspension from the deer caused hemorrhagic disease in normal white-tailed deer. Studies were conducted on the biological, physicochemical, and serologic properties of the Washington isolate. An in vitro assay system, utilizing a cultured primary of white-tailed deer fetal cells from an entire fetus, was employed for isolation and propagation of the virus. Cytopathic effect was characterized by focal development of rounded and clumped cells. Propagation was unsuccessful in suckling mice, BHK-21, and Vero cell cultures. The virus was resistant to treatment with ether, sodium deoxycholate, trypsin, oxytetracycline hydrochloride, and was sensitive to chloroform. Virus yield was not affected when infected cultures were treated with 5-iodo-2'-deoxyuridine, but dactinomycin (actinomycin D) treatment of infected cultures reduced virus yield. The virus was inactivated when heated at 70 C for 5 minutes or when exposed to pH 5 for 18 hours at 4 C. The virus was completely excluded from the filtrate by a 0.10- micronm (APD) membrane filter. Staining of infected cells with acridine orange indicated the presence of double-standard nucleic acid in the cytoplasm. Serum-neutralization tests with antiserums against the homologous virus and the New Jersey and Alberta strains of epizootic hemorrhagic disease virus resulted in neutralization of the Washington isolate. The Washington virus was not neutralized by bluetongue virus antiserum. Cells infected with the Washington isolate exhibited intracytoplasmic fluorescence by the indirect fluorescent antibody method with New Jersey and Alberta epizootic hemorrhagic disease antiserums but not with bluetongue antiserum.     

An immunodiffusion method for the identification of the species of origin of meat samples

SUMMARY The agar gel diffusion method employing suitable antiserums has been used as a convenient method for identifying species of origin of meat samples. The method can detect contamination of one species by another species at a level of between 5 to 20% depending on the sensitivity of the particular antiserum used and the species being tested, and provides a result within 24 hours.