Stable isotope analysis of sedimentary carbon in lakes can help reveal changes in terrestrial and aquatic carbon cycles. A method based on a single, photosynthetic organism, where host effects are minimised, should offer more precision than carbon isotope studies of bulk lake sediments. Here we systematically develop a method for use on fossil lacustrine diatom frustules, adapted from previous studies in marine environments. A step-wise cleaning experiment on diatomaceous lake sediments from Lake Challa, Mt, Kilimanjaro, was made to demonstrate the necessary treatment stages to remove external sedimentary carbon. Changes in soluble carbon compounds during these cleaning experiments were measured using GC/MS. Mass spectrometry methods were refined to measure the small percentage carbon of these samples and details of these methods are presented.Samples of cleaned diatoms containing <1% carbon yielded robust results. Carbon isotope analysis of diatom samples containing different species mixtures were performed and showed suggestive differences, although the effects lay within current experimental error and require further work.Unlike work on oxygen and silicon isotopes from diatom frustules, mineral contamination had no discernable impact on the diatom carbon isotope ratios from these sediments. The range of values found in the lakes investigated thus far can be interpreted with reference to the supply and nature of carbon from the catchment as well as demand generated from lake primary productivity.3