IgE and IgG antibodies in skin allergy of the horse

In horses, allergies have been characterized by clinical signs and/or intradermal (i.d.) allergen testing. Our aim was to find the first direct evidence that immunoglobulin E (IgE) mediates equine allergy. In addition, we tested the hypothesis that immediate skin reactions in horses can also be mediated by IgG. Anti-IgE affinity columns were used to purify IgE from serum of one healthy horse and three horses affected with summer eczema, an allergic dermatitis which is believed to be induced by Culicoides midges. A modified Prausnitz-Küstner experiment was performed in four clinical healthy horses by i.d. injection of the purified serum IgE antibodies. The following day, Culicoides allergen was injected at the same sites. Skin reactions were not observed in response to allergen alone, and in two horses after stimulation at any previous IgE injection site. However, the other two horses showed an immediate skin reaction at the previous injection sites of IgE obtained from allergic horses. In addition, purified monoclonal antibodies to various equine immunoglobulin isotypes were injected i.d. into six healthy horses. Immediate skin reactions were observed in response to anti-IgE (6/6 horses) and anti-IgG(T) injections (5/6 horses). The specificities of both antibodies for IgE and IgG(T), respectively, were confirmed by enzyme linked immunosorbent assays. The results provide the first direct evidence that IgE mediates classical Type-I allergy in horses and plays a major role in the pathogenesis of summer eczema. The data also suggest that IgG(T) can bind to skin mast cells and might contribute to clinical allergy.     

Practical immunoregulation: neonatal immune response variation and prophylaxis of experimental food allergy in pigs

The importance of environment in immune response is identified and the increase in prevalence of allergic, autoimmune and chronic inflammatory diseases reviewed. In particular, altered opportunity to acquire evolutionarily anticipated commensal microbiota is associated through the "hygiene hypothesis" with defective developmental and response signals to the innate and adaptive immune systems. Evidence of the detrimental effects of such environments is reviewed as is evidence for remediation using controlled exposure to bacteria or their active components such as LPS or peptidoglycan ligands for TLR and NOD-like receptors. Occurrence of major environmentally associated changes in porcine immune response phenotype are described. The prophylactic effects of heat-killed Escherichia coli given intramuscularly or of oral Lactococcus lactis on experimental ovomucoid-induced allergy in piglets are described in the context of altered immune response bias favouring reduced type-2 phenotypes. The high frequency of clinical tolerance to developing allergic signs even in the face of classical sensitization indicates possible function in this pig model of regulatory effectors such as Treg cells.     

Patch testing and allergen-specific serum IgE and IgG antibodies in the diagnosis of canine adverse food reactions

Adverse food reaction (AFR) is a common differential diagnosis for pruritic dogs. The only way to diagnose AFR is an elimination diet of 6-8 weeks with a protein and a carbohydrate source not previously fed. In humans, patch testing has been shown to be a useful tool to diagnose food allergies. In veterinary medicine, serum food allergen-specific antibody testing is widely offered to identify suitable ingredients for such diets. The aim of this study was to determine sensitivity, specificity, negative and positive predictability of patch testing with and serum antibody testing for a variety of common food stuffs. Twenty-five allergic dogs underwent an elimination diet and individual rechallenge with selected food stuffs, food patch testing and serum testing for food-antigen specific IgE and IgG. Eleven clinically normal control dogs only were subjected to patch and serum testing. The sensitivity and specificity of the patch test were 96.7 and 89.0% respectively, negative and positive predictability were 99.3 and 63.0%. For IgE and IgG the sensitivity was 6.7 and 26.7%, specificity were 91.4 and 88.3%, the negative predictive values 80.7 and 83.7% and the positive predictive values were 15.4 and 34.8%. Based on these results, a positive reaction of a dog on these tests is not very helpful, but a negative result indicates that this antigen is tolerated well. We conclude that patch testing (and to a lesser degree serum testing) can be helpful in choosing ingredients for an elimination diet in a dog with suspected AFR.     

Further purification and characterisation of horse IgE

Horse IgE was isolated from a serum pool collected from foals naturally infected with endoparasites. The serum was precipitated with ammonium sulfate, delipidated with dextran sulfate and further purified by gel filtration, anionic exchange, immunosorption or preparative polyacrylamide gelelectrophoresis. By these methods IgE could be isolated at a purity of 81%. The sera from rabbits immunized with the purified horse serum fractions were tested using reversed passive cutaneous anaphylaxis and an enzyme linked immunosorbent assay (ELISA). By the ELISA method cross reaction of rabbit anti horse IgE sera to human, mouse and rat myeloma IgE was demonstrated. Rat myeloma IgE also served to monitor the production of antibodies to horse IgE in rabbits.     

Gastrointestinal aspects of food allergy: A review

Food allergy in dogs and cats, although rare, is a well recognised cause of skin disease. However, despite reports of conditions responding to dietary manipulation, evidence for gastrointestinal manifestations of food allergy is scarce. Difficulties involved in reaching a clinical diagnosis of food allergy are discussed in an attempt to explain this enigma. The distinction between food allergy and intolerance, the complexity of the gastrointestinal immune system and the limited understanding of the mechanisms by which food allergy develops are reviewed.     

Induced and spontaneous IgE antibodies to Dermatophagoides farinae in dogs and cats: evidence of functional heterogeneity of IgE

Enzyme-linked immunosorbent assays (ELISAs) were developed to measure IgE antibodies specific for Dermatophagoides farinae in dogs and cats. Although higher levels were detected in atopic dogs and cats than in normal animals without skin disease, the differences were not statistically significant. On the other hand, levels in dogs and cats that were reared under laboratory conditions, and thus presumably not exposed to house dust mites, were either very low or undetectable. IgE antibodies were induced in 10 laboratory-reared cats using low-dose antigenic stimulation in aluminium hydroxide. All cats developed detectable IgE, but not all developed positive skin tests. However, serum from those cats with positive skin tests were able to give positive Prausnitz–Küstner (PK) tests. The canine data, together with previous work on basophil histamine release, suggests that the distinction between atopic and normal dogs may result from a heterogeneity of either IgE or of the high-affinity mast cell receptor. The feline data can only be explained by the existence of a heterogeneity of IgE.     

Anti-neosporal IgG and IgE antibodies in canine neosporosis

Neospora caninum infection provokes neurological disorders, recurrent abortion and death in dogs and cattle. Dogs are both intermediate and definitive host of N. caninum. Thus, the development of sensitive and specific immunoassays to diagnose canine neosporosis is essential to control this disease. This work investigated serum anti-neosporal IgG and IgE antibodies in 140 dogs represented by 30 healthy animals (group I), 11 dogs showing acute N. caninum infection (group II), 50 urban dogs with serological evidence of canine neosporosis in indirect fluorescent antibody test (IFAT) (group III) and 49 urban dogs without clinical and laboratory evidences of neosporosis (group IV). Enzyme-linked immunosorbent assay (ELISA) and western immunoblotting, both using a soluble N. caninum tachyzoite antigen (SNA), investigated these two isotypes of antibodies, while a Urea-ELISA measured the avidity of the IgG antibodies. Anti-Toxoplasma gondii IgG antibodies were also investigated in the animals. Anti-neosporal IgG was found in all animals from groups II and III, whereas 32.7% (16/49) of dogs from group IV were reactive. IgG antibodies of low avidity were demonstrated in dogs from group II (median 35.3%), while animals from groups III and IV had IgG antibodies of high avidity (medians of 61.5% and 61.7% respectively). IgE antibodies were found in four (13.3%) and five (16.6%) dogs from groups III and IV respectively. Dogs presenting acute infection (group II) or chronic infection (group III) had IgG antibodies to several neosporal antigens, mainly of 29-30 and 35 kDa, while 13 of 16 dogs from group IV recognized antigens from 14 to 170 kDa. Antibodies to T. gondii were detected in 36 of 50 (72%) sera from group III and 25 of 49 (51%) sera from group IV. We concluded that IgG-ELISA and Urea-ELISA with SNA may substitute for IFAT in both laboratory routine and epidemiological studies of canine neosporosis.     

Monoclonal anti-IgE antibodies in the diagnosis of dog allergy

The availability of anti-dog IgE monoclonal antibodies has enabled development of highly specific ELISA assays for measuring antigen-specific IgE in dog serum. In this article the authors propose criteria for evaluation of these monoclonals and demonstrate that some anti-human IgE monoclonals recognize dog IgE. Combinations of two or more monoclonal antibodies can enhance assay sensitivity; for example, a mixture of DE38.HRPO and 4F4.HRPO conjugates detect total dog IgE in the range 10–10000 ng/mL. Results are reported from nine clinical studies conducted in Europe, Japan and Australia involving more than 400 dogs in which serologic IgE determinations performed using the CMG IMMUNODOT strip system for house dust mites, storage mites, flea, grass pollens and moulds were compared with immediate skin test findings. House dust mites were identified as the common major dog allergen throughout these areas although regional reactions to food allergens were observed. These results confirm that the CMG IMMUNODOT system is a valuable and reliable diagnostic test for dog allergy.     

人工感染绦虫条件下猪血清中免疫球蛋白E的分离提取与鉴定

注射疫苗可以引起猪发生过敏反应,尤其品种猪发病率较高,治疗不及时极易导致动物死亡。过敏反应主要由免疫球蛋白E(immunoglobulin E,IgE)介导发生,分离纯化到猪体内的IgE是研究猪过敏反应ELISA检测方法的前提和基础。IgE在正常人血清中浓度极低,正常猪血清中IgE含量还未见有确定报道。本试验研究的目的是通过人工感染方法对感染绦虫试验猪血清样品中的IgE进行提取,并且通过SDS-PAGE电泳对血清中提取的IgE进行鉴定,从而建立猪血清IgE的最佳提取方法。     

Development of nasal, fecal and serum isotype-specific antibodies in calves challenged with bovine coronavirus or rotavirus

Unsuckled specific pathogen free calves were inoculated at 3-4 weeks of age, either intranasally (IN) or orally (O) with bovine coronavirus or O plus IN (O/IN) or O with bovine rotavirus. Shedding of virus in nasal or fecal samples, and virus-infected nasal epithelial cells were detected using immunofluorescent staining (IF), ELISA or immune electron microscopy (IEM). Isotype-specific antibody titers in sera, nasal and fecal samples were determined by ELISA. Calves inoculated with coronavirus shed virus in feces and virus was detected in nasal epithelial cells. Nasal shedding persisted longer in IN-inoculated calves than in O-inoculated calves and longer than fecal shedding in both IN and O-inoculated calves. Diarrhea occurred in all calves, but there were no signs of respiratory disease. Calves inoculated with rotavirus had similar patterns of diarrhea and fecal shedding, but generally of shorter duration than in coronavirus-inoculated calves. No nasal shedding of rotavirus was detected. Peak IgM antibody responses, in most calves, were detected in fecal and nasal speciments at 7-10 days post-exposure (DPE), preceeding peak IgA responses which occurred at 10-14 DPE. The nasal antibody responses occurred in all virus-inoculated calves even in the absence of nasal shedding of virus in rotavirus-inoculated calves. Calves inoculated with coronavirus had higher titers of IgM and IgA antibodies in fecal and nasal samples than rotavirus-inoculated calves. In most inoculated calves, maximal titers of IgM or IgA antibodies correlated with the cessation of fecal or nasal virus shedding. A similar sequence of appearance of IgM and IgA antibodies occurred in serum, but IgA antibodies persisted for a shorter period than in fecal or nasal samples. Serum IgG1 antibody responses generally preceeded IgG2 responses and were predominant in most calves after 14-21 DPE.     

猪伪狂犬病的防控及净化措施

猪伪狂犬病是目前规模化生猪养殖生产中一种危害较为严重且难以防控的疫病,该病在临床生产中传染性强,死亡率高,且易形成隐性感染,很难将病原彻底清除,给生猪养殖业带来很大的经济损失.本文在猪伪狂犬病原相关生物学特性的基础上,根据猪伪狂犬病的临床症状及病理变化,对该病进行系统诊断及流行情况分析,并根据实际生产情况提出了针对性的...     

Allergic pulmonary and ocular tissue responses in the absence of serum IgE antibodies (IgE) in an allergic dog model

Allergen-specific serum IgE may be insensitive as a marker for IgE-mediated reactions at the mucosal level. Five of six atopic beagle dogs developed high ovalbumin (OVA)-specific serum IgE levels after sensitization. This study aimed to show that these dogs still express allergen-specific IgE at the pulmonary and ocular mucosal levels and in the skin even when corresponding serum IgE was below the detection limit. When serum IgE levels were negative, all dogs exhibited allergic reactions at the tissue level. Specifically, they displayed positive ocular reactions after an ocular OVA challenge. After airway challenge with aerosolized OVA, five out of six animals reacted with decreased compliance and increased resistance of the lungs. Furthermore, an eosinophilia in the bronchoalveolar lavage fluid (BALF) was observed. Four weeks after the last exposure to OVA, IgE-positive BALF cells were seen in all animals. Six weeks on, all dogs still displayed positive skin reactions to OVA. This indicates that not only skin testing but also detection of ocular and pulmonary allergic tissue reactions including cell-bound IgE in BALF can serve as more sensitive and lasting surrogate markers of hypersensitivity in the allergic dog model than detection of allergen-specific serum IgE levels.     

The use of monoclonal antibodies in an enzyme immunospot assay to detect isotype-specific antibody-secreting cells in pigs and chickens

Monoclonal antibodies directed against porcine immunoglobulin isotypes G, G1, G2, M, and A and against chicken immunoglobulin isotopes G, M, and A were tested in an antigen-specific spot-forming cell (SFC) assay based on the principle of the enzyme immunoassay. The SFC assay was used to quantitate ovalbumin (OA)-specific antibody-secreting cells (ASC) in pigs that had been primed and boosted with OA. The SFC assay was also used to quantitate trinitrophenyl (TNP)-specific ASC in chickens that had been primed with TNP-conjugated keyhole lympet haemocyanin (TNP-KLH). Although, the classical plaque-forming cell (PFC) assay cannot reliably detect isotope-specific ASC in pigs and chickens, it can detect these cells in mice. Therefore, we compared the OA- and TNP-specific SFC assays with PFC assays that were specific for these antigens in mice. The study demonstrated that the SFC assay is superior to the PFC assay in detecting both OA-specific ASC and TNP-specific ASC. The frequencies of OA-specific and TNP-specific SFC detected in mice were of the same order of magnitude as those detected in pigs and chickens. We concluded that the SFC assay is the better method for quantitating ASC in pigs, chickens, and probably all domestic animals for which isotype-specific monoclonal antibodies are available.     

Comparison of intradermal testing and serum testing for allergen-specific IgE using monoclonal IgE antibodies in 84 atopic dogs

OBJECTIVE: To compare an ELISA measuring serum allergen-specific IgE with intradermal skin testing in canine atopic dermatitis. PROCEDURE: Eighty-four dogs with the clinical diagnosis of atopic dermatitis underwent intradermal skin testing and serum testing for allergen-specific IgE. Tests were performed in a blinded fashion. Positive reactions were compared and the sensitivity and specificity of the serum test (using intradermal skin test as the standard) were determined overall and for individual allergen groups (grass pollens, weed pollens, tree pollens, house dust mites and fleas). RESULTS: The sensitivity of the ELISA overall was 90.4%. Evaluating the individual allergen groups, the sensitivity for dust mite hypersensitivity was 95.1%, for fleas 85.4%, for tree pollens 84.3%, for grass pollens 95.1% and for weed pollens 96.4%. The specificity was 91.6% overall, for dust mites 96.3%, for fleas 92.7%, for tree pollens 95.2%, for grass pollens 94% and for weed pollens 80.7%. CONCLUSION: The evaluated ELISA seemed reliable for the diagnosis of atopy in practice and can be recommended as a screening test prior to intradermal skin testing or for use in dogs when immunotherapy is not a therapeutic option.     

Analysis of the expression and secretion of isotypes of sheep B cell immunoglobulins with a panel of isotype-specific monoclonal antibodies

Monoclonal antibodies to sheep light chain, IgM and IgG were produced and used to assess total immunoglobulin (Ig) synthesis by sheep B cells in culture and antibodies to specific antigens. By using these antibodies in a dual fluorescence-activated cell sorting analysis of sheep efferent lymph B lymphocytes the percentage change in surface Ig isotype of B lymphoblasts from IgM to IgG after the antigenic stimulation of the local lymph node was measured. An extension of this analysis to paired blood and afferent or efferent lymph B cells made it possible to investigate the recirculation characteristics of B cells expressing different Ig isotypes.     

Gastrointestinal food allergy and its role in large domestic animals

The significance of food allergy as a primary cause for gastrointestinal disturbances in domestic animals, especially calves and piglets, is discussed. The immunological backgrounds and pathogenesis are described in some detail. The clinical and pathological manifestations in animals are related to those in man. Diagnostic possibilities, therapy and prevention, as far as known in animals, are mentioned and, based on human experiences, further extensions are proposed.     

IgE and IgG antibodies to flea antigen in differing dog populations

A radioimmunoassay was developed for the detection of IgG and IgE canine antibodies against partially purified flea antigen. Low background levels were found in flea naive dogs, but high levels of both IgE and IgG antibodies were found in many sera from dogs with clinical flea hypersensitivity. In sera from non-allergic dogs exposed chronically to fleas, IgE levels differed little from background, and levels of IgG anti-flea antibodies were much lower than those from the flea allergic group. The results suggest that chronic flea exposure may result in partial or complete tolerance rather than hyposensitization in the commonly accepted sense.     

Monoclonal anti-equine IgE antibodies with specificity for different epitopes on the immunoglobulin heavy chain of native IgE

In this study we describe the generation of monoclonal antibodies (mAbs), which recognize different epitopes of the equine IgE constant heavy chain. Equi-murine recombinant IgE (rIgE), composed of the murine V(H)186.2 heavy chain variable region, linked to the equine IgE constant heavy chain and expressed together with the murine lambda(1) chain in J558L cells was used to immunize BALB/C mice. A total of 17 different mAbs were obtained, which recognized the rIgE heavy chain constant region. None of the mAbs reacted with monoclonal equine isotypes IgM, IgG1 (IgGa), IgG3 (IgG(T)), IgG4 (IgGb) or isolated equine light chains, IgGc and IgA from horse serum, or the native mAb B1-8delta, expressing the same heavy chain variable regions and light chains. One of the mAbs (alphaIgE-132) recognized the recombinant equine IgE, but did not recognize any protein in equine serum, i.e. native IgE. A total of 16 mAbs detected a serum protein of approximately 210,000Da on Western blots, corresponding to the expected MW of native IgE. In addition, one of the mAbs (alphaIgE-176) detected a protein of 76,000Da under reducing conditions, most likely the equine IgE heavy chain. According to binding inhibition studies, the equine IgE specific mAbs recognize at least two different epitopes of the equine IgE. In an ELISA using two anti-IgE mAbs which recognized different epitopes, no significant differences in the concentration of total serum IgE could be detected between adult Icelandic horses with IgE-mediated type I allergy (summer eczema) and healthy control animals. In Icelandic horse foals, no serum IgE could be measured 6 months post partum. All anti-IgE mAbs recognized a small population (1.3+/-0.5%) of leukocytes from adult Icelandic horses by surface immunofluorescence, but no cells could be detected in foal blood. The stained leukocytes from adult horses could be enriched by magnetic cell sorting and contained 32% basophils, 53% monocytes and/or large lymphocytes, 13% small lymphocytes and 2% eosinophils.