Autofluorescence spectroscopy and imaging have been studied for the early detection and classification of (pre)malignancies of the oral mucosa. In the present review we will give an overview of the literature on autofluorescence imaging and spectroscopy for various clinical questions. From the studies performed so far we hope to conclude whether autofluorescence spectroscopy and imaging are helpful in the diagnosis of lesions of the oral mucosa, and if this is the case: for which clinical questions they are suitable. A strong emphasis is put on in vivo human studies of the oral mucosa.