Objectives: Day/night variations in cellularity, percentage of CD4+, CD8+ and double-positive (CD4+–CD8+) lymphocytes, lipopolysaccharide (LPS)- and concanavalin A (Con A)-induced lymphocyte proliferation and natural killer (NK) activity, and the effect of timed administration of recombinant human prolactin (h-PRL) on the above-mentioned parameters were investigated in the submaxillary lymph nodes and spleen of adult male mice. Results: In controls, the percentage of CD4+, double-positive lymphocytes, LPS- or Con A-induced blastogenic proliferation and NK activity in the spleen differ during the dark phase as compared to the light phase. When administered during the dark period, h-PRL induced immunosuppresion in the percentage of CD4+, double-positive (CD4+–CD8+) lymphocytes. Con A- and LPS-induced lymphocyte proliferation and NK activity as compared to untreated controls. When h-PRL was administered during the light period, the cellularity increased, and h-PRL was immunosuppressive in Con A- and LPS-induced lymphcoyte proliferation and NK activity as compared to controls. Moreover, in control submaxillary lymph nodes the cellularity, percentage of CD8+, double-positive lymphocytes, blastogenic proliferation in the presence of Con A and LPS and NK activity differ when comparing the dark with the light phase. When administered during the dark period h-PRL induced immunosuppression in the percentage of double-positive (CD4+–CD8+) lymphocytes, Con A- and LPS-induced lymphocyte proliferation as compared to controls. When h-PRL is administered during the light period, no effects were observed. Conclusions: These results indicate the existence of differential day/night variations in the cellular immune response depending upon the lymphoid organ considered. Because of the administration of h-PRL a differential modulation of this circadian variation was also observed.