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A double immunofluorescent technique that permitted the nuclear localization of both the oestrogen receptor and the Ki-67 antigen (marker of cell proliferation) within the same tissue section was developed and used to determine the relationship between the oestrogen receptor and proliferation within the major zones of the primate endometrium. Endometrial tissue was obtained from ovariectomized rhesus monkeys in which the hormonal pattern of oestradiol and progesterone was simulated with Silastic implants containing each hormone. The oestrogen receptor was detected using the H222 rat monoclonal antibody and Texas Red conjugated streptavidin. Immunofluorescent staining of a mouse monoclonal antibody to the Ki-67 antigen was detected by an antibody to mouse IgG conjugated with fluorescein isothiocyanate. Primary antibodies were co-incubated with tissue sections followed by sequential detection of the oestrogen receptor and Ki-67. A dual wavelength filter was used to monitor simultaneously both immunofluorescent patterns. Application of this technique to study regulation of endometrial proliferation and oestrogen receptor content by progesterone showed that the endometrium exhibited two different responses to progesterone: luminal and glandular epithelia and stromal cells in the functionalis and zone III of the basalis showed coincident decreases of oestrogen receptor and proliferation, whereas the oestrogen receptor in glandular epithelia of zone IV (basalis) was not downregulated and there was an increase in proliferation.

reproduction

A double immunofluorescent method for simultaneous analysis of progesterone-dependent changes in proliferation and the oestrogen receptor in endometrium of rhesus monkeys