Barrett’s oesophagus (BO) is a complicated condition at the gastroesophageal junction in which normal squamous epithelium is changed to columnar and leads to oesophageal adenocarcinoma (OA). In the past decades, the prevalence of Barrett’s disease and mortality rate of adenocarcinoma has significantly increased throughout the word. Data has shown that molecular pathogenesis of disease has not been clearly identified. However, a wide-range and successful administration of probiotics in cancer and gastrointestinal diseases has lead to the investigation into the possible inhibitory role of probiotics in oesophageal cancer. This study was conducted to evaluate the inhibitory effect of probiotics on the expression of biomarkers in an in vitro model. Two different Barrett’s oesophageal cell lines were selected to co-culture with B. longum and Lactobacillus acidophilus to measure expression of IL-18, TNFα, p53 (tumour suppressor gene), cyclooxygenase 2 and CDX1 (caudal type homeobox 1) genes. In addition, two different aspects of probiotic administration, therapeutic and prophylactic test were also examined. Results showed that micro-organisms could inhibit expression of biomarkers and therapeutic culture conditions were more effective than prophylactic tests. The results obtained suggest that it is possible to incorporate the administration of probiotics in BO and OA prevention.