Quantitation of Anaplasma marginale major surface protein (MSP)1a and MSP2 epitope-specific CD4+ T lymphocytes using bovine DRB3*1101 and DRB3*1201 tetramers

Immunogenetics. 2006 Sep;58(9):726-39. doi: 10.1007/s00251-006-0140-3. Epub 2006 Aug 19.


Antigen-specific CD4+ T cells play a critical role in protective immunity to many infectious pathogens. Although the antigen-specific CD4+ T cells can be measured by functional assays such as proliferation or cytokine enzyme-linked immunospot, such assays are limited to a specific function and cannot quantify anergic or suppressed T cells. In contrast, major histocompatiblity complex (MHC) class II tetramers can enumerate epitope-specific CD4+ T cells independent of function. In this paper, we report the construction of bovine leukocyte antigen MHC class II tetramers using a novel mammalian cell system to express soluble class II DRA/DRB3 molecules and defined immunodominant peptide epitopes of Anaplasma marginale major surface proteins (MSPs). Phycoerythrin-labeled tetramers were either loaded with exogenous peptide or constructed with the peptide epitope linked to the N terminus of the DRB3 chain. A DRB3*1101 tetramer loaded with MSP1a peptide F2-5B (ARSVLETLAGHVDALG) and DRB3*1201 tetramers loaded with MSP1a peptide F2-1-1b (GEGYATYLAQAFA) or MSP2 peptide P16-7 (NFAYFGGELGVRFAF) specifically stained antigen-specific CD4+ T cell lines and clones. Tetramers constructed with the T-cell epitope linked to the DRB3 chain were slightly better at labeling CD4+ T cells. In one cell line, the number of tetramer-positive T cells increased to approximately 94% of the CD4+ T cells after culture for 21 weeks with specific antigen. This novel technology should be useful to track the fate of antigen-specific CD4+ T-cell responses in cattle after immunization or infection with persistent pathogens, such as A. marginale, that modulate the host immune response.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Anaplasma marginale / genetics
  • Anaplasma marginale / immunology*
  • Anaplasma marginale / pathogenicity
  • Animals
  • Antigens, Bacterial / genetics
  • Antigens, Bacterial / immunology*
  • Bacterial Outer Membrane Proteins / genetics
  • Bacterial Outer Membrane Proteins / immunology*
  • Base Sequence
  • CD4-Positive T-Lymphocytes / immunology*
  • Cattle
  • Cell Line
  • DNA / genetics
  • Erythrocytes / immunology
  • Erythrocytes / microbiology
  • Genes, MHC Class II
  • Histocompatibility Antigens Class II / chemistry*
  • Histocompatibility Antigens Class II / genetics
  • Immunodominant Epitopes / genetics
  • In Vitro Techniques
  • Molecular Sequence Data
  • Protein Structure, Quaternary
  • Transfection


  • Antigens, Bacterial
  • Bacterial Outer Membrane Proteins
  • BoLA-DRB3 antigen
  • Histocompatibility Antigens Class II
  • Immunodominant Epitopes
  • major surface protein 1a, Anaplasma
  • msp2 protein, Anaplasma marginale
  • DNA