In vitro stimulation of bovine milk lymphocytes standardization of the assay for bovine brucellosis

L. F. Ayanwale, J. B. Kaneene, D. W. Johnson, Charles C Muscoplat, R. K. Anderson

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Lymphocytes of bovine milk origin were investigated by immunostimulation in vitro to standardize the assay for measuring the immune responses of the cells which might be useful in further understanding the immunopathology and diagnosis of bovine brucellosis. The lymphocytes were separated from whole freshly collected milk by centrifugation. The pellet of lymphocytes was washed in RPMI-1640 medium, cultured at different concentrations for different days and with Brucella abortus soluble antigen strain 1119-3 and Concanavalin A. Each culture was labelled with 1.0 μCi of methyl-[3H]thymidine 16-18 hours prior to termination of incubation at 37 C. Termination was done by cooling to 4 C. The cells were harvested for liquid scintillation counting spectrometry. In the groups of calfhood vaccinated cows and nonexposed milkers, a milk lymphocyte concentration of 2.0 × 106/ml of medium yielded a statistically significant blastogenesis. The Brucella abortus soluble antigen concentration of 4.4 μg of protein/well was found optimal to induce significant immunostimulation. A period of 4 days of incubation of the milk lymphocyte in the test was found optimal in inducing statistically significant blastogenesis in this system.

Original languageEnglish (US)
Pages (from-to)343-352
Number of pages10
JournalComparative Immunology, Microbiology and Infectious Diseases
Issue number3-4
StatePublished - 1981


  • Lymphocyte stimulation
  • bovine brucellosis


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