1. A new technique has been developed for making serial measurements of water and solute absorption from the lumen of isolated small intestine.2. The isolated intestine is perfused in a single pass with a segmented flow of slugs of liquid separated by bubbles of oxygen-carbon dioxide mixture. Simultaneous collections are made of effluent from the lumen and of the fluid which is transported across the mucosa. This latter fluid appears to be a fair sample of the tissue fluid.3. Conditions in the lumen can be changed within less than 5 min. The effects of two or more treatments applied to the same segment of intestine can be determined and the time course of a change in luminal conditions.4. The rate of appearance of solutes on the serosal side depends on the rate of water absorption, and changes exponentially towards a steady state. The rate constant is a function of tissue fluid volume.5. In the steady state the concentration of glucose in the tissue fluid is 71 mM when the luminal concentration is 28 mM, and is 45 mM when the luminal concentration is 8.3 mM.6. For solutes such as glucose for which reflux from tissue fluid to lumen is small relative to flux from lumen to tissue fluid, the time of attainment of a steady state in secretion is usually 50-60 min.7. For solutes such as sodium for which the reflux is relatively high, the steady state may be reached in 15-20 min.8. The K(m) for glucose absorption (14-19 mM) is much lower than is found with unsegmented flow perfusion.9. These findings emphasize problems in interpreting results from other types of intestinal preparation.10. The rate of glucose absorption from the lumen falls only gradually when the luminal sodium concentration is reduced abruptly. In contrast the rate of glucose absorption falls suddenly when the luminal glucose concentration is reduced abruptly. This suggests that glucose absorption is not directly dependent on luminal sodium ions.