The fluorescence activated cell sorter (FACS) offers a new approach to purification and analysis of pituitary cell types. Using dispersed rat anterior pituitary cells 29.5 ± 3.2% of pituitary cells can be recovered from the FACS of which up to 93% are viable. More importantly, these cells actively adhere to cell culture dishes and respond to both the gonadotropin releasing hormone (GnRH) by increasing LH secretion and to dopamine by decreasing prolactin secretion. Using the narrow angle light scatter discriminator combined with the detection of LH beta subunit antibody conjugated to highly fluorescent microspheres, immunocytochemically identified gonadotropes have been enriched from the normal control range of 7.4 ± 1.4% to 52.3 ± 11% of sorted cells (n = 4). This enhancement was also confirmed by measuring the ratio of LH to prolactin in the cell homogenates of control and sorted groups. This ratio rose from 8 to 650, 26 to 35, and 55 to 170 in three independent experiments. The ability of the fluorescent microspheres bound to the LH beta subunit antibody to identify gonadotropes was also suggested by studies in which pituitary cells were coincubated with both this “fluorescein” antibody probe and “rhodamine” microspheres which had been conjugated to D-Lys6GnRH. Only 5% of the cells bound more than five spheres and, in each case, these cells bound both types of spheres. Thus it is likely that these were gonadotropes. The use of multichannel fluorescence detection should allow the simultaneous use of several different fluorescent probes for greater accuracy of selection of cell types (e.g., use of microspheres conjugated to antibody directed against the secretory product and other microspheres with different fluorescent characteristics conjugated to hypophysiotropic hormones). It appears that FACS technology will be readily applicable to a variety of endocrine tissues for separation and analysis of specific cell types and for preparation of enriched populations of these cell types in a viable form.