Hepatic extraction, cellular and subcellular localization of gelatin stabilized Tc-99m sulfur colloid was studied in the rat model with time sequenced microautoradiography from 15 min to 24 h following I.V. administration of the tracer. Hepatic lobular and cellular distribution, quality and quantity of focal grain pattern, grain clusters, remained essentially constant for the period of study. Grain clusters were associated predominantly with Kupffer cells lining the peripheral segment of hepatic lobular sinusoids. Subcellular localization of gelatinized Tc sulfur colloid, stained prior to the I.V. administration with osmium tetroxide, was demonstrated with a transmission electron microscope in unosmicated liver tissue. Extracted Tc sulfur colloid particles were attached in groups to cytoplasmatic membranous intrasinusoidal projections of activated Kupffer cells. Intracytoplasmatic phagocytosis was not demonstrated. The kinetic arrest and en groupe extraction of Tc sulfur colloid particles at the Kupffer cell membrane suggests a specific membrane receptor site and specific Tc sulfur colloid particle-plasma protein interaction at the time of extraction. Hepatic extraction of gelatinized Tc sulfur colloid thus reflects primarily extra and intra hepatic hemodynamics and does not serve as an indicator of phagocytic hepatic reticuloendothelial system function.