Bacillus Calmette Guerin (BCG) intravesical therapy of bladder cancer is arguably the most effective immunotherapy for any human solid tumour. It combines a high incidence of remission induction with a low level of side effects and a low rate of recurrence. Its mechanism, however, remains poorly understood. In this study we have investigated whether gamma delta T lymphocytes, which are known to be activated by mycobacteria, are preferentially induced in patients urine following therapy. This has necessitated the development of a procedure which facilitates the preservation, enrichment and detection of small number of lymphocytes [especially of those bearing gamma delta T cell receptor (TCR)I which appear in patients urine. Here we describe in detail a method for phenotyping of a minor subpopulation of lymphocytes in patients' urine, namely, gamma delta T lymphocytes which comprised less than 0.1% of all urinary sells. Using this technique we have found gamma delta T cells in the urine of all patients. Furthermore, the patients could be separated into 2 distinct groups, with low and high numbers of gamma delta cells (0.5-5% and 5-20% respectively of the CD3 positive cells). The elevation of gamma delta T cells was observed locally but not in peripheral blood and the detected gamma delta T were almost entirely of the V delta 2 gamma 9 subset.

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