Faecal proteomics is ideally suited to interrogate gastrointestinal tract (GI tract) pathologies, giving an alternative biological sample to blood for the discovery, validation and quantitative assay of specific biomarkers for disease detection, monitoring and surveillance. As the stool passes through the GI tract it “samples” the local environment, collecting proteins arising from leakage, secretion or exfoliation, whilst also sampling the gut microbiome, known to play an important role in health and disease. Importantly potential biomarkers may be present at relatively elevated levels in faeces compared with serum, facilitating detection of low abundance proteins such as carcinoembryonic antigen (CEA). Collection is non-invasive, requires no unpleasant bowel preparation, and can be undertaken at home using a simple collection kit without the need for trained staff or a formal health care visit. Protocols will be presented for the discovery and validation of potential biomarkers/biomarker panels for colorectal cancer (CRC) and other GI tract pathologies, illustrating the potential to quantitatively measure low-level faecal components such as CEA. A targeted MRM approach to detect known disease-associated proteins will also be presented. Finally the potential of faecal proteomics to support faecal transplantation, that is showing exceptional potential for the treatment of C. difficile infection, will also be discussed