Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME) is a physically and mentally disabling illness with immunological abnormalities. Dysfunction in the innate and adaptive cells in particular Natural Killer cells (NK) and regulatory T cells (Tregs) have been identified in CFS/ME patients. However, an expansive analysis of T and B cells has not been conducted in severely affected CFS/ME patients in comparison to those who are moderately affected. The objective of this study was to analyse adaptive immune cells, including Tregs, gamma delta T (γδ T), iNKT and B cells in severe and moderately affected CFS/ME patients to assess potential immunological dysregulation. CFS/ME patients were examined using the 1994 Centres for Disease Control and Prevention Case Definition and the International Consensus Criteria for ME. Health, mobility and quality of life questionnaires were utilised to distinguish CFS/ME participants as moderately or severely affected and to confirm non-fatigued controls. Severe CFS/ME patients were housebound visited in their homes to ensure wellbeing. Whole blood samples collected from patients and controls (n=65) were labelled with a combination of monoclonal antibodies, following which assessment of T cells, Tregs, gamma delta T, iNKT and B cells phenotypes were performed on the flow cytometer. Data confirmed differences in adaptive immune cells in severe and moderate CFS/ME patients compared to the non-fatigued control group. These results are suggestive of adaptive immune cell perturbations, potentially related to the immune dysregulation demonstrated in moderate and severe CFS/ME and contributing to the overall immune dysfunction observed in the illness.