Hydrocarbons and organic biomarkers extracted from black shales and other carbonaceous sedimentary rocks are valuable sources of information on the biodiversity and environment of early Earth. However, many Precambrian hydrocarbons including biomarkers are suspected of being younger contamination. An alternative approach is to study biomarkers trapped in oil-bearing fluid inclusions by bulk crushing samples and subsequently analysing the extracted hydrocarbons with gas chromatography-mass spectrometry. However, this method does not constrain the hydrocarbons to one particular oil inclusion, which means that if several different generations of oil inclusions are present in the sample, a mix of the content from these oil inclusions will be analysed. In addition, samples with few and/or small inclusions are often below the detection limit. Recently, we showed that it is possible to detect organic biomarkers in single oil-bearing fluid inclusions using time-of-flight secondary ion mass spectrometry (ToF-SIMS). In the present study, single fluid inclusion analysis has been performed on Proterozoic samples for the first time. Four individual oil-bearing fluid inclusions, found in 1430. Ma sandstone from the Roper Superbasin in Northern Australia, were analysed with ToF-SIMS. The ToF-SIMS spectra of the oil in the different inclusions are very similar to each other and are consistent with the presence of n-alkanes/branched alkanes, monocyclic alkanes, bicyclic alkanes, aromatic hydrocarbons, and tetracyclic and pentacyclic hydrocarbons. These results are in agreement with those obtained from bulk crushing of inclusions trapped in the same samples. The capability to analyse the hydrocarbon and biomarker composition of single oil-bearing fluid inclusions is a major breakthrough, as it opens up a way of obtaining molecular compositional data on ancient oils without the ambiguity of the origin of these hydrocarbons. Additionally, this finding suggests that it will be possible to analyse minute oil samples beyond the capability of established techniques. This may allow the biomarker record of the biosphere, as preserved in fluid inclusions, to be extended further back in time, and hence makes it possible to more accurately trace the early evolution of life on Earth, and search for life on other planets or moons.