Correlative confocal and transmission electron microscopy

CD11c-YFP mice were perfused with PBS and SMG were fixed in situ by left ventricle injection of 1.5% glutaraldehyde/2% paraformaldehyde in 0.1 M sodium cacodylate buffer (pH 7.4). SMG were harvested and immersed in the same solution for 16 hours. Fixed samples were cryoprotected in 30% sucrose before embedding in optimum cutting temperature and freezing. Thirty-micrometer sections were processed for confocal imaging. After confocal image acquisition, coverslips were gently removed, and sections adherent to the slide were processed for TEM as previously described (64).