Individual antigens of cattle. Monoclonal antibodies reacting with bovine B cells (BoWC3, BoWC4 and BoWC5).
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Biomedical subjects
C J Howard
Publications and source records attributed to C J Howard.
Competitive binding with putative Bo5 (CD5) cluster of monoclonal antibodies.
The relationship of seven monoclonal antibodies, putatively to the Bo5 (CD5) antigen, was tested. Five of the mAbs were confirmed to be directed against the Bo5 antigen. Three mAbs, CC29, BLT-1 and 8C11, effectively blocked binding to bovine PBM of mAb CC17, previously reported to be directed against this antigen. MAb 8-3F4 also blocked binding of mAb CC17, but less effectively than the others. MAbs IL-A67 and 79-5 did not inhibit binding of mAb CC17 because of antibody allelic specificity or technical reasons.
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Investigating monoclonal antibodies to bovine "null" cell antigens using two-colour immunofluorescence.
Eighteen monoclonal antibodies (mAbs) putatively to non T4/T8 (null) cell antigens were tested by two-colour immunofluorescence and antibody binding inhibition (blocking), with one selected mAb (CC15) that previous studies had indicated to be specific for null cells. None of the other mAbs blocked binding of CC15 to lymphocytes. Three main patterns of reaction were observed in two-colour immunofluorescence studies: mAbs that stained the same cells as CC15, mAbs that only stained a sub-population of the cells that stained with CC15 and mAbs that stained a sub-population of the cells that stained with CC15 but also some cells that did not react with CC15.
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Analysis of the reactivity of anti-bovine CD8 monoclonal antibodies with cloned T cell lines and mouse L-cells transfected with bovine CD8.
Mouse L-cells transfected with bovine CD8 and two Theileria parva-infected cloned T cell lines expressing bovine CD8 were used to screen the panel of ten monoclonal antibodies (mAbs) submitted to the workshop. Eight of the ten mAbs reacted with the transfectant and both the cloned T cell lines. However, two mAbs CC58 and BAT82A did not recognise the transfectant and only reacted with one of the T cell lines. Further biochemical studies indicated that the eight mAbs react with both homo- and heterodimeric forms of bovine CD8 whilst the two mAbs CC58 and BAT82A react with only heterodimeric forms. These data suggest that bovine DC8 is encoded by two genes as is the case in mouse and man.
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Immunohistology of workshop monoclonal antibodies to the bovine homologue of CD1.
Six monoclonal antibodies putatively to the BoCD1 antigen were compared by immunohistology on cryostat sections from a range of tissues. The different staining patterns observed allowed the mAbs to be placed in three groups (a) 20-27, (b) CC13, CC14, TH97A and (c) CC20, CC40. An ovine mAb VPM5 did not stain bovine tissues sufficiently strongly to enable a comparison with the other CD1 mAbs.
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Polymorphism of the CD4 and CD5 differentiation antigens in cattle.
Monoclonal antibodies (mAbs) specific for bovine CD4 and CD5 antigens have been found to identify polymorphic determinants on these molecules. In the case of CD5, mAb IL-A67 recognises one allotypic form of the antigen while four other CD5-specific mAbs in the workshop (CC17, CC29, BLT-1 and 8C11) recognise a second allotype. The CD4-specific mAbs submitted to the workshop reacted with the cells of all animals tested. However, a further two mAbs (CC26 and IL-A18) specific for CD4 were found to react with cells only from about 85% of animals tested. Sequential immuno-precipitation experiments together with family studies showed that the allotypes of CD4 and CD5 are both inherited in a simple Mendelian manner and are co-dominantly expressed. One of the CD5 allotypes was not detected in Bos taurus animals while the gene frequency of the second allotype was only about 10% in the B. indicus animals tested. The gene frequency of the CD4 allotype detected by CC26 and IL-A18 was similar in the two sub-species.
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Identification of a bovine surface antigen uniquely expressed on CD4-CD8- T cell receptor gamma/delta+ T lymphocytes.
In this study, two monoclonal antibodies, IL-A29 and CC15, are described that identify a novel bovine cell surface marker of 215/300 kDa. The antibodies reacted with a discrete population of resting lymphocytes in peripheral blood which, in young animals, constituted about 25% of the mononuclear cells. Thymus, lymph nodes and spleen contained less than 5% positive cells. These cells were negative for surface Ig, a monocyte/granulocyte marker, and the T lymphocyte antigens CD2, CD6, CD4 and CD8. Immunohistological analyses revealed the presence of IL-A29/CC15-positive lymphocytes in the thymic medulla, in the outer cortex of lymph nodes, in the marginal zones of the spleen, in the dermal and epidermal layers of the skin and in the lamina propria of the gut. The IL-A29/CC15+ cells in unfractionated blood mononuclear cells responded in autologous and allogeneic mixed lymphocyte cultures, and when purified they responded to concanavalin A in the presence of recombinant interleukin 2. These observations suggested this population of cells belonged to the T cell lineage. In order to unambiguously define their lineage, cDNA clones encoding bovine T cell receptor (TcR) and CD3 proteins were isolated. Northern blot analyses of IL-A29/CC15+ cell populations and of established cell lines of various lineages demonstrated that they expressed TcR delta and CD3 gamma, delta and epsilon mRNA: TcR alpha was not expressed, whereas only a truncated form of TcR beta mRNA was present. These results indicate that the IL-A29 and CC15 antibodies define a unique population of CD4-CD8-, gamma/delta T cells.
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Immunological responses to bovine virus diarrhoea virus infections.
Infection of normal calves with bovine virus diarrhoea virus (BVDV) is a transient self-limiting infection that can result in a period of immunosuppression. The virus appears to be able to replicate in all of the major lymphocyte sub-populations as well as in accessory cells. This may result in the leukopenia that is often a sequel of infection and affects B-cells as well as the T-cell sub-populations expressing either BoCD4 or BoCD8 antigens. B and T-cell responses are affected as a consequence of exposure to BVDV and there is a reduced ability to control other infections. Evidence is summarised and shows that immunoglobulin is an important mediator of immunity to infection with BVDV. Although the foetus can mount an immune response in the latter part of gestation, during the first trimester it does not. A specific state of tolerance is induced and this is associated with change in the proportion of certain lymphocyte sub-populations and ability to respond to immune stimulation.
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Characterization of a bovine leucocyte differentiation antigen of 145,000 MW restricted to B lymphocytes.
A new bovine B-cell differentiation antigen is described that is detected by three monoclonal antibodies (mAb). The antigen is not an immunoglobulin and is precipitated from peripheral B cells as a molecule with an approximate molecular weight (MW) of 120,000 or 145,000 before and after reduction, respectively. Data obtained from two-colour cytofluorimetry and immunohistochemistry confirmed that the antigen was found only on mature B cells and on cells with dendritic morphology in the follicles of the organized lymphoid tissues. Its level of expression is directly correlated with that of IgM on peripheral blood B cells and Theileria parva-transformed B cells. The marker was also expressed on the peripheral cells which expressed surface IgG. Based on the antigen's cellular distribution, biochemistry and histochemistry, it is considered to be analogous to the human CD21 antigen.
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In vivo depletion of BoT4 (CD4) and of non-T4/T8 lymphocyte subsets in cattle with monoclonal antibodies.
The effect on certain immune responses of depleting two distinct lymphocyte subpopulations in vivo by inoculating calves with monoclonal antibodies (mAb) was examined. An mAb directed against the BoT4 antigen (the bovine homologue of CD4) effectively removed the BoT4+ lymphocytes from peripheral blood mononuclear cells (PBM). Compared to controls, treated calves showed a reduced antibody response to human O red blood cells and to ovalbumin. PBM prepared from BoT4-depleted animals also had a significantly reduced ability to respond in vitro to the mitogens phytohemagglutinin, concanavalin A and pokeweed mitogen. An mAb directed against a second numerically large bovine lymphocyte subpopulation i.e. BoT2-, BoT4-, BoT8- (CD2-, CD4-, CD8-), that may be homologous to the CD4-, CD8- cells in man and rodents that synthesize the gamma/delta+ T cell receptor, was also used for in vivo depletion. Compared to controls, calves depleted of this subpopulation showed an enhanced antibody response. The proliferative response of PBM to pokeweed mitogen was also significantly increased but responses to concanavalin A and phytohemagglutinin remained unchanged. The results suggest this lymphocyte subpopulation has a nonspecific suppressor activity acting on B cell responses either directly or through an effect on T helper cells. The non-T4/T8 cells are found extensively in the epithelium and lamina propria of the mucosa of the alimentary tract but not in T cell areas of the lymph nodes, tonsil and spleen. These non-T4/T8 cells may thus be, or contain, an intraepithelial lymphocyte population with a suppressor function.
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Protection against respiratory infection with bovine virus diarrhoea virus by passively acquired antibody.
Susceptibility to infection with bovine virus diarrhoea virus (BVDV) was compared for calves with varying amounts of specific antibody in their sera passively acquired from the ingestion of colostrum. Challenge consisted of intranasal exposure to a strain of BVDV isolated from an outbreak of respiratory disease. Resistance to infection, as judged by nasopharyngeal shedding of virus, was directly related to the titre of neutralizing antibodies in sera. Besides protecting against infection of the upper respiratory tract, passive antibody, which was mainly IgG1, also protected against viraemia and, to a lesser extent, leukopenia. In the presence of colostral antibody, neutralizing and IgG1 antibody responses were apparently inhibited, but a specific IgG2 response occurred.
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Demonstration of two allelic forms of the bovine T cell antigen Bo5 (CD5) and studies of their inheritance.
Two monoclonal antibodies (mAbs), CC17 and IL-A67, which are specific for the bovine equivalent of the CD5 antigen, Bo5, were each found to react with the cells of some animals but not others. The cattle tested were all positive for one or both of the mAbs, but the level of expression on cells expressing both determinants was slightly lower than that on cells expressing either of the determinants on their own. Both mAbs precipitated an antigen of 67 kD. However, sequential immunoprecipitation experiments with cells that reacted with both mAbs demonstrated that the determinants are present on two different sets of molecules. These findings suggested that the mAbs recognize two co-dominantly expressed allelic forms of Bo5. This was confirmed in family studies, with groups of full- and half-sibling offspring of sires and dams of defined phenotypes. These experiments also showed that the gene encoding the Bo5 antigen is not linked to the major histocompatibility complex (MHC). The frequencies of the two alleles, which have been designated Bo5.1 and Bo5.2, in the cattle populations tested were 100% and 0%, respectively, in Bos taurus, and 10% and 90%, respectively, in Bos indicus.
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Investigation of bovine gut associated lymphoid tissue (GALT) using monoclonal antibodies against bovine lymphocytes.
Gut associated lymphoid tissue of the small and large intestine of calves and cows has been compared morphologically and quantitatively using monoclonal antibodies to bovine lymphocytes. B cells were significantly decreased in the ileum of the cow compared to the calf. Significantly increased numbers of T cells were present in cell suspensions of all lymphoid areas of the cow compared to the calf. T lymphocyte subsets were quantified into cryostat sections of lymphoid tissues expressing BoT4, and BoT8 antigens demonstrated increased numbers in follicular and dome areas of the discrete Peyer's patches of the small and large intestine of the cow. BoT4+, BoT8+, and the non-BoT4/BoT8+ T cell subsets were increased in the mucosa of the cow as compared to the calf. Similarities in structure and lymphocyte composition of the discrete Peyer's patches of the small intestine, cecum and colon and isolated single follicles in the large intestine suggest similar functional properties.
Analysis of Variance↗
Experimental infection of cattle in early pregnancy with a cytopathic strain of bovine virus diarrhoea virus.
Nine pregnant heifers, in early gestation (63 to 107 days), were infected intranasally or in utero with cytopathic bovine virus diarrhoea virus (BVDV) and each dam seroconverted. All nine calves developed to full term; four were stillborn, of which one had seroconverted but virus was not recovered from their tissues. One of the five liveborn calves appeared to have seroconverted in utero to an adventitious BVDV infection in late pregnancy but the remaining four were not viraemic and showed a normal secondary antibody response to BVDV infection at about six months old. Thus, in contrast to results with noncytopathic virus there was no evidence that infection in utero with cytopathic virus could result in a persistent viraemia or immunotolerance. It is suggested that cells able to support a persistent viraemia with cytopathic virus may not be developed in the young fetus.
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Two monoclonal antibodies (CC17, CC29) recognizing an antigen (Bo5) on bovine T lymphocytes, analogous to human CD5.
Two new monoclonal antibodies (CC17 and CC29) raised against bovine thymocytes are described. The antibodies, both of which were IgG1, recognize a molecule of approximately 67,000 molecular weight on bovine T cells. They react T cells in peripheral blood, the lymph node paracortex and the periateriolar lymphoid sheath in the spleen. Both the cortex and medulla of the thymus are stained but the medulla reacts more intensely. They do not stain B cells in peripheral blood, the ileal Peyer's patch, the cortex or the primary follicles in lymph nodes. No activity was found on cells outside the lymphoid system, i.e. monocytes, alveolar macrophages or endothelial and epithelial tissue. The antigen recognized is considered to be the bovine homologue of CD5 (T1) in humans and Lyt1 in mice. The mAbs appear to be particularly useful for detecting cells in the peripheral blood of young calves which are of the T cell lineage but do not express BoT2 or the mature pan T cell antigen recognized by mAb IL-A27 and may thus allow identification of a population of bovine lymphocytes previously described as null cells.
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Characterization of a bovine thymic differentiation antigen analogous to CD1 in the human.
Three monoclonal antibodies (MoAb), TH97A, CC13, and CC14, define a thymic differentiation antigen in cattle. The antigen is expressed on 50-60% of bovine thymocytes, located mainly in the cortical areas, but is not expressed on peripheral blood mononuclear cells (PBMC). In cryostat sections of lymph node, the antibodies react with large dendritic-like cells in the paracortical regions. They also react with a proportion of the large 'frilly' cells in afferent lymph and with dendritic-like cells in the dermis. The antibodies apparently do not react with cells in the epidermis. Biochemical analysis of the antigen recognized by MoAb TH97A reveals two bands of 44 kDa and 12 kDa under reducing conditions. These polypeptides are distinct from bovine class I major histocompatibility complex molecules reactive with the MoAb w6/32. The tissue distribution of positive cells together with results of biochemical analyses indicate that the antigen recognized by these MoAb is the bovine analogue of the human CD1.
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Protection against respiratory disease in calves induced by vaccines containing respiratory syncytial virus, parainfluenza type 3 virus, Mycoplasma bovis and M dispar.
A field trial to assess the ability of two vaccines to protect calves against respiratory disease was carried out on a large beef rearing unit in southern England over the two winters of 1983 to 1984 and 1984 to 1985. A quadrivalent vaccine containing the killed antigens of respiratory syncytial virus, parainfluenza virus type 3, Mycoplasma bovis and M dispar or a vaccine containing only the respiratory syncytial virus component were inoculated into 246 and 245 calves, respectively; 245 calves remained as unvaccinated controls. The calves were reared in seven batches and outbreaks of disease occurred in five; significant protection was achieved in the four batches in which disease was associated with respiratory syncytial virus and M bovis infection, together or independently. The death rate from pneumonia was 9 per cent in the control group, 2 per cent in the calves inoculated with the quadrivalent vaccine (P less than 0.001), a protection rate of 77 per cent, and 3 per cent in the calves inoculated with the respiratory syncytial virus vaccine (P less than 0.01), a protection rate of 68 per cent. The proportion of calves receiving treatment for respiratory disease was 38 per cent in the control group, 25 per cent in the calves inoculated with the quadrivalent vaccine (P less than 0.001) and 27 per cent in the calves inoculated with the respiratory syncytial virus vaccine (P less than 0.01). The results show that protection against respiratory disease can be achieved by parenteral vaccination of calves with the appropriate inactivated microorganisms.
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Field trial of a quadrivalent vaccine against calf respiratory disease.
A quadrivalent vaccine containing the killed antigens of respiratory syncytial virus, parainfluenza virus type 3, Mycoplasma dispar and M bovis, emulsified with an oil adjuvant, was tested for efficacy against naturally occurring calf respiratory disease. Three batches of beef cattle aged 12, seven and three weeks at the time of first vaccination were used. Within each batch of approximately 100 animals, half were vaccinated subcutaneously on three occasions, three weeks apart and half served as unvaccinated controls. Over the trial period, from November 1981 to May 1982, 27 per cent of the control calves were treated for respiratory disease compared with 16.3 per cent of the vaccinated animals. This reduction of non-fatal disease in the vaccinated animals represented a protection rate of almost 40 per cent and was statistically significant (P less than 0.05). Mortality was also reduced from 3.4 per cent in the control calves to 1.9 per cent in the vaccinated animals but this difference was not statistically significant. During a major outbreak of disease associated with respiratory syncytial virus, the protection rate increased to 69 per cent (P less than 0.01). Furthermore, in the batch of cattle aged seven weeks at first vaccination there was significantly less pneumonic consolidation at death in the vaccinated animals than in the control animals (P less than 0.05).
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