Gamma Delta (γδ) T lymphocytes contain the unique capability of responding to pathogens in both an innate and acquired immune response. Previously, γδ lymphocytes have been reported to respond to Mycobacteria tuberculosis determined by proliferation and IFN-γ production. Unlike alpha beta (αβ) lymphocytes, γδ lymphocytes constitutively express a natural killer receptor providing γδ lymphocytes the capability for innate cytolytic functions. A new cytolytic assay by flow cytometry was reported capable of determining natural killer activity using K562 cells as targets without the need for radioactive materials. The objectives of this study were to first apply the flow cytometer-based assay to assess γδ lymphocytes natural killer activity following animal vaccination with Mycobacterium bovis Bacillus Calmette-Guérin (BCG). Secondly, to optimize the flow cytometer assay in order to detect antigen specific cytolytic activity to mycobacterium and to compare the cytolytic activity of γδ lymphocytes to CD-8 lymphocytes. γδ lymphocytes increased in NK activity (P=0.012) following animal vaccination with M. bovis BCG. Both innate (P=0.02) and acquired antigen-specific cytolytic activity (P=0.04) increased following incubation with M. bovis-infected monocytes. In conclusion, flow cytometric-based assay is a sensitive and reliable tool to determine cytolytic activity of γδ T-lymphocytes against mycobacterium.
Bibliographical noteFunding Information:
Special thanks to Dr. Jack Risdal, Nextran for the donation of the animals used in this study; Dr. Bruce Walcheck and Shelia Alexander for helping with the setup of FL-4 laser on the flow cytometry and supplying us with the K562 cell line. Michael Olin was supported by National Institute of Health, National Research Service Award T32 DA07097 from the National Institute on Drug Abuse.
- Flow cytometer
- Mycobacterium bovis
- Mycobacterium tuberculosis
- Natural killer cytolytic activity
- γδ lymphocytes