Quantifying in vivo recruitment of naïve CTL towards individual dendritic cells by standard immunofluorescence histology

Dendritic cells (DCs) attract naïve cytotoxic CD8+ T lymphocytes (CTL) by chemokines, in order to activate them for immune defense against viruses and tumors. Here we describe a protocol for quantifying such recruitment towards individual DCs in secondary lymphatics by standard immunofluorescence histology. We used reporter mice expressing green fluorescent protein under a chemokine promoter and adoptively transferred into them CTL labeled with a red fluorochrome. Tissue sections from these animals were stained with a third color identifying anatomical regions, in our case the splenic B cell zone, which encircles the T cell zone in which DCs and CTL are located. The number of CTL in that zone and their location adjacent to chemokine-producing CTL can be determined by computer-assisted digital analysis