The clinical utility of cross-linked tetrameric hemoglobin solutions is limited by peripheral vasoconstriction thought to be due to scavenging of nitric oxide. In addition, transfusion of crude preparations of hemoglobin polymers can cause arterial hypertension. We tested the hypothesis that eliminating low-molecular-weight components from the polymer solution would prevent extravasation and its associated pressor response. A zero-link polymer of bovine hemoglobin was developed without chemical linkers left between the tetramers. Transfusion of unprocessed preparations of these polymers in rats resulted in appearance of the polymer in the renal hilar lymph. However, eliminating the low-molecular-weight components with a 300-kDa diafiltration resulted in an average hydrodynamic radius of 250 Å and in undetectable levels of polymer in hilar lymph. Exchange transfusion in anesthetized rats and cats and in awake cats produced no increase in arterial pressure. In anesthetized cats, exchange transfusion with an albumin solution reduced hematocrit from 30 to 18%, increased cerebral blood flow, and dilated pial arterioles. In contrast, reducing hematocrit by transfusing the diafiltered polymer did not increase cerebral blood flow as pial arterioles constricted. These results are consistent with the hypothesis that the increase in arterial pressure associated with cell-free hemoglobin transfusion depends on hemoglobin extravasation. Constriction observed in the cerebrovascular bed with a nonextravasating hemoglobin polymer at low hematocrit is presumably a regulatory response to prevent overoxygenation at low blood viscosity.