Albuminuria is a key marker of renal injury and a major risk factor for cardiovascular disease. In vivo imaging techniques with fluorescent albumin have allowed visualization of its movement within the whole kidney but they could not distinguish between intact and degraded albumin. To visualize albumin degradation in proximal tubular cells in vivo we used an albumin conjugate (dye quenched (DQ)-albumin), which only fluoresces when it is degraded. In cultured proximal tubule cells, the fluorescent signal from DQ-albumin was dependent on endocytosis and lysosomal function and showed that at any time about 40% of endocytosed DQ-albumin was degraded. Significant accumulation of conventional Texas Red-labeled albumin and degraded DQ-albumin was found in rat proximal tubules 5 min after injection. Importantly, no hint of DQ-albumin was detected in the serum, suggesting that the fluorescent signal in the proximal tubules was derived from tubular degradation of intact albumin. Our study shows that DQ-albumin, together with conventional fluorescent conjugates of intact albumin, can be used to visualize albumin degradation by proximal tubules in vivo.