The buccal micronucleus cytome assay has previously been used and validated using visual scoring microscopy to measure distinct differences between the cytome profiles associated with normal ageing relative to that for premature ageing clinical outcomes such as Down’s syndrome and Alzheimer’s disease. The aim was to develop a laser scanning cytometry (LSC) protocol to automatically score the frequency of the most abundant cell types in exfoliated buccal cell samples, and the nuclear DNA and neutral lipid content within each cell type. Cells derived from the buccal mucosa were applied to a microscope slide, stained with Fast Green (cytoplasm), Oil Red O (neutral lipid) and DAPI (nuclei). The frequency of the different buccal cell types was scored in exfoliated buccal cell samples collected in a minimally invasive manner from healthy elderly subjects aged from 58 to 88 years old with result showing that (i) on average the frequencies of basal, transitional, differentiated and karyolytic cells were 16%, 35%, 24%, 25% respectively and (ii) the DNA content and neutral lipid content of basal cells was significantly higher compared with that of other cell types. This automated LSC method, although not corresponding precisely with visual scoring methods, provides a non-subjective and rapid approach that allows for the co-location of neutral lipids and DNA content within the specific cell types and can be combined with other cellular markers for high content automated analyses in population studies.