016 Driving diagnostic change; microchimerism in thymus transplant patients

Athymic patients are now receiving thymus transplants, allowing for mature T-cells to begin populating the bloodstream. Although much care is taken when preparing the thymus tissue for transplantation to ensure the complete removal of donor-derived T cells, the patients show clinical symptoms, such as rashes. These symptoms, usually classed as autoimmunity, may be explained by the presence of donor T cells in the recipient and may, therefore, be linked to alloimmunity rather than autoimmunity. Current methods (short tandem repeat analysis) for chimerism analysis are sensitive to 99%, however, the presence of extremely low levels of donor cells may fall below this threshold and avoid detection. In order to overcome this and detect microchimerism at very low levels, we are developing a method to increase sensitivity up to 1:100,000 cells, based on qPCR targeting sequence polymorphisms (SP). We used a panel of markers to screen recipient (pre-transplant) and donor samples for their specific SP. This allows us to identify suitable markers to discriminate between the recipient and donor T cells in a post-thymic transplant sample. The standard curve was created by using healthy human blood to create a serial 10-fold dilution in order to accurately quantify the results. Preliminary results show that the donor thymus, although washed out, still contains donor T cells at very low levels. This implies that a number of symptoms may be caused by alloimmunity rather than autoimmunity. Using this qPCR based method for the detection of microchimerism, we have shown in a small number of patients tested so far that we can accurately quantify the amount of donor T cells present in the recipient, meaning appropriate treatment options can be given to reduce symptoms.