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Microvascular damage, often resulting in renal failure, is a common complication of diabetes. Transcapillary fluorescein escape rate (TCFER) as monitored by intravital microscopy has been used as an indicator of the extent of capillary damage in diabetes and to assess improvement in microvascular function after combined kidney-pancreas transplant. However, fluorescein anion binds to plasma albumin, and albumin-ligand binding may be altered in the presence of renal disease. The purpose of this study was to compare fluorescein binding by plasma from diabetics with renal failure with plasma from healthy nondiabetics. Fluorescein binding by plasma from seven type I diabetics awaiting kidney-pancreas transplant and seven healthy adults of similar age and sex was studied using ultrafiltration and dialysis. There was no significant difference in the apparent albumin binding of fluorescein at physiologically relevant fluorescein concentrations, even though the TCFER was significantly increased in the diabetics as compared with the controls. Hippurate, a ligand that accumulates in renal failure, did alter fluorescein binding in a defatted albumin solution but not sufficiently to account for the differences in TCFERs. These data indicate that impaired albumin binding of fluorescein does not contribute significantly to the TCFER in diabetics with renal failure.