In this study, we evaluated the effect of splenectomy on the febrile response to lipopolysaccharide (LPS) and on its clearance by Kupffer cells (KC), its main uptake tissue and presumptive source of endogenous pyrogens. Conscious splenectomized (Splex) or sham-operated (Sham) guinea pigs were challenged with high and low doses of LPS, administered iv (0.05 or 2 µg/kg) or ip (2 or 8 µg/kg) 7 and 30 days after the surgery. Using fluorescent microscopy, FITC-LPS uptake (75 µg/Kg, iv or ip) was assessed at 15, 30, and 60 min after injection. LPS iv at 0.05 µg/kg did not evoke fever in the Sham controls, but caused a ~1.2 °C Tc rise in the Splex guinea pigs. LPS iv at 2 µg/kg induced a ~1.8 °C greater rise in the Tcs of the Splex guinea pigs than of their controls; their post-febrile recovery also stabilized at a higher level. LPS ip at 2 and 8 µg/kg 7 days post-surgery induced ~1.4 and ~1.8 °C higher fevers in the Splex animals, respectively, but with the same latencies, than in their Sham counterparts. LPS ip at 2 and 8 µg/kg 30 days post-surgery also increased the febrile responses of the asplenic animals by ~1.6 and ~1.8 °C, respectively. FITC-LPS at 7 days was detected in the controls as abundant granular fluorescent patches within presumptive KC in the liver sinusoids 15 min after its administration; the label density was reduced at 30 min and almost 0 at 60 min. In the Splex animals, in contrast, the labeling was significantly denser and remained unchanged through all three time points; this effect was still present 30 days after surgery. Similar results were obtained at 60 min after FITC-LPS ip injection. Gadolinium chloride pretreatment (-3 days) of the Splex animals significantly reduced both their febrile responses to LPS ip (8 µg/kg) and their KC uptake of FITC-LPS 7 days post surgery. Thus, splenectomy increases the magnitude of the febrile response of guinea pigs and the uptake of systemically administered LPS, suggesting that the spleen may modulate endotoxic fever, perhaps by influencing the avidity of KC for LPS.