The intrathymic location of cells responding to the thymocyte growth peptide (TGP) was investigated in guinea pigs. Thymus cells were labelled by topical application of a fluorescein isothiocyanate solution, and the percentage of fluorescent cells analyzed by flow cytometry. Large cells forming rosettes with rabbit erythrocytes, known to respond to TGP, had a lower percentage of fluorescent cells than non-rosetting cells, which do not respond to TGP. If the TGP-responding cells are anatomically segregated in the guinea pig thymus, this might indicate that they are preferentially located in the deep, juxtamedullary part of the cortex, where one major proliferating compartment is situated.