Biochemical evidence of the expression of two major histocompatibility complex class I genes on bovine peripheral blood mononuclear cells

Joosten, Irma; Teale, A.J.; Poel, A. van der; Hensen, E. J.

doi:10.1111/j.1365-2052.1992.tb00030.x

Cited by 24 publications

(2 citation statements)

References 23 publications

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“…It has been reported that N-terminus of β 2 m is involved in the formation of the W6/32 epitope (Shields and Ribaudo 1998) and in the case presented here, the N-terminal methionine of the recombinant bovine met-β 2 m likely interferes with W6/32 detection. Alternatively, the failed W6/32 recognition may be explained by the lack of pan-specificity of the antibody to BoLA-I when associated with bovine met-β 2 m (Joosten et al 1992). Substituting bovine met-β 2 m with human β 2 m, lacking the N-terminal methionine, however, proved highly suitable for the W6/32 based LOCI detection with BoLA-6*01301.…”

Section: Resultsmentioning

confidence: 99%

Characterization of binding specificities of bovine leucocyte class I molecules: impacts for rational epitope discovery

et al. 2014

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The binding of peptides to classical major histocompatibility complex (MHC) class I proteins is the single most selective step in antigen presentation. However, the peptide-binding specificity of cattle MHC (bovine leucocyte antigen, BoLA) class I (BoLA-I) molecules remains poorly characterized. Here, we demonstrate how a combination of high-throughput assays using positional scanning combinatorial peptide libraries, peptide dissociation, and peptide-binding affinity binding measurements can be combined with bioinformatics to effectively characterize the functionality of BoLA-I molecules. Using this strategy, we characterized eight BoLA-I molecules, and found the peptide specificity to resemble that of human MHC-I molecules with primary anchors most often at P2 and P9, and occasional auxiliary P1/P3/P5/P6 anchors. We analyzed nine reported CTL epitopes from Theileria parva, and in eight cases, stable and high affinity binding was confirmed. A set of peptides were tested for binding affinity to the eight BoLA proteins and used to refine the predictors of peptide–MHC binding NetMHC and NetMHCpan. The inclusion of BoLA-specific peptide-binding data led to a significant improvement in prediction accuracy for reported T. parva CTL epitopes. For reported CTL epitopes with weak or no predicted binding, these refined prediction methods suggested presence of nested minimal epitopes with high-predicted binding affinity. The enhanced affinity of the alternative peptides was in all cases confirmed experimentally. This study demonstrates how biochemical high-throughput assays combined with immunoinformatics can be used to characterize the peptide-binding motifs of BoLA-I molecules, boosting performance of MHC peptide-binding prediction methods, and empowering rational epitope discovery in cattle.

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Section: Resultsmentioning

confidence: 99%

Characterization of binding specificities of bovine leucocyte class I molecules: impacts for rational epitope discovery

et al. 2014

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“…The vital nucleic acid stain Sytox Orange, which penetrates cells with compromised plasma membranes but will not cross uncompromised cell membranes, was added at 5 m (final) and incubated (10 min) prior to automated reading of results by twochannel fluorescence microscopy (Cell Observer, Carl Zeiss, x5, ZEN 2012 software). Positive (pan-MHC I-specific mAb; IL-A88; [28]) and negative (FCS) controls were included with each serum-cell combination, together with a complement control (no serum) and live cell control (no serum and heat-inactivated complement). Total cells (∼4000/sample) and dead cells were counted using a macro in Image-J software [29].…”

Section: Complement-dependent Cytotoxicity Assaymentioning

confidence: 99%

Haematopoietic depletion in vaccine-induced neonatal pancytopenia depends on both the titre and specificity of alloantibody and levels of MHC I expression

Bell

MacHugh

Connelley

et al. 2015

Vaccine

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Bovine Neonatal Pancytopenia (BNP) is a disease of calves characterised by haematopoietic depletion, mediated by ingestion of alloantibodies in colostrum. It has been linked epidemiologically to vaccination of the dams of affected calves with a particular vaccine (Pregsure) containing a novel adjuvant. Evidence suggests that BNP-alloantibodies are directed against MHC I molecules, induced by contaminant bovine cellular material from Madin-Darby Bovine Kidney (MDBK) cells used in the vaccine's production. We aimed to investigate the specificity of BNP-alloantibody for bovine MHC I alleles, particularly those expressed by MDBK cells, and whether depletion of particular cell types is due to differential MHC I expression levels. A complement-mediated cytotoxicity assay was used to assess functional serum alloantibody titres in BNP-dams, Pregsure-vaccinated dams with healthy calves, cows vaccinated with an alternative product and unvaccinated controls. Alloantibody specificity was investigated using transfected mouse lines expressing the individual MHC I alleles identified from MDBK cells and MHC I-defined bovine leukocyte lines. All BNP-dams and 50% of Pregsure-vaccinated cows were shown to have MDBK-MHC I specific alloantibodies, which cross-reacted to varying degrees with other MHC I genotypes. MHC I expression levels on different blood cell types, assessed by flow cytometry, were found to correlate with levels of alloantibody-mediated damage in vitro and in vivo. Alloantibody-killed bone marrow cells were shown to express higher levels of MHC I than undamaged cells. The results provide evidence that MHC I-specific alloantibodies play a dominant role in the pathogenesis of BNP. Haematopoietic depletion was shown to be dependent on the titre and specificity of alloantibody produced by individual cows and the density of surface MHC I expression by different cell types. Collectively, the results support the hypothesis that MHC I molecules originating from MDBK cells used in vaccine production, coupled with a powerful adjuvant, are responsible for the generation of pathogenic alloantibodies.

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The evolution of MHC class I genes in cattle

Ellis

Holmes

Staines

et al. 2000

Major Histocompatibility Complex

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Biochemical evidence of the expression of two major histocompatibility complex class I genes on bovine peripheral blood mononuclear cells

Abstract: 113 114 1. Joosten et al.expressed on normal bovine PBMC. In IEF analysis the additional use of mAb recognizing polymorphic determinants on serologically defined A-locus products highly facilitated the detection and typing of second locus products.

Cited by 24 publications

References 23 publications

Characterization of binding specificities of bovine leucocyte class I molecules: impacts for rational epitope discovery

Characterization of binding specificities of bovine leucocyte class I molecules: impacts for rational epitope discovery

Haematopoietic depletion in vaccine-induced neonatal pancytopenia depends on both the titre and specificity of alloantibody and levels of MHC I expression

The evolution of MHC class I genes in cattle

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