Advanced glycosylation end‐products (AGEs) are reported to be a risk factor for cardiovascular mortality in hemodialysis (HD) patients. As serum AGEs can change with dialysis, measurement of AGEs deposited in the skin by autofluorescence (SAF) is now a recognized method of measuring AGEs. An arteriovenous fistula (AVF) is the preferred way to access blood in HD patients, and as the creation of an AVF changes blood flow distribution in the arm, we wished to determine whether this affected SAF deposition in the skin. SAF was measured using the AGE reader, which directs ultraviolet light at an intensity range of 300‐420 nm (peak 370 nm) in the arms of HD patients dialyzing with an AVF. We measured SAF in 267 patients, 60.3% male, 46.1% diabetic, median duration of dialysis 34.7 (15.1‐64.2) months with AVF. The median SAF was lower in the AVF arm (median 3.4 (2.9‐4.2) vs. 3.7 (3.2‐4.5) AU, P < .001), and for the 160 patients with an upper arm AVF (3.5 (2.9‐4.3) vs. 3.8 (3.2‐4.5) AU, P < .001), but not for the 107 patients dialyzing with a forearm AVF ((3.4 (2.8‐4.2) vs. 3.6 (3.0‐4.5) AU, P = .085). Blood flow was greater for upper arm AVF compared to forearm AVFs (1190 (770‐1960) vs. (930 (653‐1250) mL/min, P = .007), but there was no association between blood flow and SAF (P > .05). AVF alters blood flow in the arm, and we found that SAF measurements were lower in the arm with AVF. We suggest that SAF measurements are made in the non‐AVF arm.