A protocol to detect apoptotic dendritic cells in murine lymph nodes using multiphoton microscopy

This protocol describes a method to detect and measure the percentage of apoptotic or live dendritic cells (DCs) in popliteal lymph nodes (PLNs). DCs labelled with fluorescent cell trackers are subcutaneously (s.c.) injected in mice. After allowing the DCs to reach the PLNs, animals are intravenously injected with FLIVOTM, a permeant fluorescent reagent that selectively marks apoptotic cells. Of note, the fluorophore moiety of the latter reagent should be selected so as its emission wavelength can be distinguished under the confocal microscope from that of the fluorescent cell tracker used to label the cells. Explanted PLN are then examined under a two-photon microscope to analyse for the presence of apoptotic cells among the DCs injected.