Three-dimensional confocal thermal imaging using anti-Stokes luminescence

High-resolution thermal images of dye-stained live cells are obtained using confocal anti-Stokes luminescence on a confocal laser scanning microscope. Stokes and anti-Stokes fluorescence signal ratio imaging, using dual laser excitation with a fast alternate line scan, is used to monitor dynamical temperature changes at submicrometer resolution. Temperature mapping of a dye-doped polymer film with local heating is shown as an example. Three-dimensional mapping of local temperature changes in living cells induced by an external laser as the heating source, is obtained with temperature resolution better than 0.5 K.