Detection and Characterizationof Cellular Immune Responses Using Peptide–MHC Microarrays
Figure 4
Detection of a Weak Immune Response to Vaccination
CTLs from OVA-vaccinated and mock-vaccinated mice were analyzed in parallel using an MHC array and flow cytometry analysis. Mice received base-of-tail injections of OVA/Freund's adjuvant emulsions or PBS (mock)/Freund's adjuvant on day 0 and day 7. Draining lymph nodes were harvested on day 11, and dissociated into a single-cell suspension.
(A) CTLs were enriched on an anti-CD8-bead column. Cells (2 × 106 and 3.2 × 106) from the OVA- and mock-vaccinated mice, respectively, were incubated on identical but separate arrays printed with OVA/Kb tetramer, LCMV/Kd (control) tetramer, and various antibodies. The three panels show the relevant array results. A rare population of cells from the OVA-vaccinated mouse was captured on the OVA/Kb (left), but not on the LCMV/Kd (right), tetramer spot. The cells captured on the OVA/Kb tetramer spot were CD8+ (as determined by counter-staining using an anti-CD8–FITC mAb on the array). An arrowhead points to some of the cells that were bound to that spot. The cells from the mock-vaccinated mouse did not bind the OVA tetramer (middle spot) or the LCMV/Kd tetramer (data not shown). Spot regions are marked with a blue color by overlaying the tetramer's PE fluorescent signal onto the DIC image.
(B) FACS analysis of the cells from the OVA-vaccinated (left panel) and mock-vaccinated mice (right). Of the total CD8+ cells from the vaccinated mouse, 0.27% co-stain with OVA/Kb, compared with 0.01% in the mock-vaccinated mouse.