Tracking and imaging immune cells in vivo non-invasively would offer insights into the immune responses induced by vaccination. Here we report a cancer vaccine consisting of polymer-coated NaErF₄/NaYF₄ core–shell down-conversion nanoparticles emitting luminescence in the near-infrared spectral window IIb (1,500–1,700 nm in wavelength) and with surface-conjugated antigen (ovalbumin) and electrostatically complexed adjuvant (class-B cytosine–phosphate–guanine). Whole-body wide-field imaging of the subcutaneously injected vaccine in tumour-bearing mice revealed rapid migration of the nanoparticles to lymph nodes through lymphatic vessels, with two doses of the vaccine leading to the complete eradication of pre-existing tumours and to the prophylactic inhibition of tumour growth. The abundance of antigen-specific CD8⁺ T lymphocytes in the tumour microenvironment correlated with vaccine efficacy, as we show via continuous-wave imaging and lifetime imaging of two intravenously injected near-infrared-emitting probes (CD8⁺-T-cell-targeted NaYbF₄/NaYF₄ nanoparticles and H-2K^b/ovalbumin_257-264 tetramer/PbS/CdS quantum dots) excited at different wavelengths, and by volumetrically visualizing the three nanoparticles via light-sheet microscopy with structured illumination. Nanoparticle-based vaccines and imaging probes emitting infrared light may facilitate the design and optimization of immunotherapies.