We examined the topographic cell affinity of different types of hormone-producing cells in the rat anterior pituitary gland. Pituitaries were removed from 8-10 week-old male SD rats, and were fixed in sublimate-formalin. The 40 μm sections were obtained with a Microslicer, then double-immunolabeled (FITC and TexasRed methods) using antibodies against five types of pituitary hormones. Topographic affinity was examined three-dimensionally using a confocal microscope, and was expressed as attachment rate. Statistical analysis revealed that the different types of hormone-producing cells did not exist at random locations in the gland, but rather showed specific topographic affinities. The following pairs of cell types were observed to have significantly high affinities: LH and PRL cells, ACTH and GH cells, and TSH and GH cells. In contrast, the following pairs of cell types were observed to have no significant affinities: GH and PRL cells, ACTH and LH cells, TSH and LH cells, ACTH and TSH cells, and cells of the same type. These specific topographic affinities between different types of anterior pituitary cells may be intimately related to cell-to-cell interactions in the gland.