The diffusive in vitro uptake of homologous 125I-albumin (MA, nmol.cntdot.cm-2) and Evans blue dye (EBD) (ME, nmol.cntdot.cm-2) by the deendothelialized canine aorta from serum and from a simple albumin solution with and without EBD and with and without vigorous stirring was measured in 18 preparations. MA and ME were significantly smaller from serum than from a simple albumin solution; vigorous stirring of the liquid phase caused a slight decrease (.apprx. 5%) in MA and increase (.apprx. 9%) in MH; MA was not influenced by the presence of EBD; and at least 90% of the radioactivity in the tissue was free 125I-albumin with an electrophoretic mobility identical to its nonlabeled cohort molecules and albumin in the original reagent. These observations confirm the identity of the tissue radioactivity with the labeled protein in the reagent, show that < 10% of the labeled protein is irreversibly bound in the tissue, indicate that significant concentration gradients do not occur in the reagent phase, and indicate that albumin appears to interact with other plasma components in the reagent phase.