To assess the relative bioavailability of bis-glycinate bound Cu, 90 Angus-cross steers (265 ± 21 kg) were blocked by body weight (BW) to pens with GrowSafe bunks and randomly assigned to dietary treatments (14 to 18 steers/treatment): 0 mg supplemental Cu/kg dry matter (DM; CON), 5 or 10 mg supplemental Cu/kg DM as Cu sulfate (CS5; CS10) or bis-glycinate bound Cu (GLY5; GLY10). Steers received a high antagonist growing diet (analyzed 4.9 mg Cu/kg DM, 0.48% S, and 5.3 mg Mo/kg DM). Steers were weighed at the beginning (days 1 and 0) and end (days 125 and 126) of the trial to determine average daily gain (ADG) and gain:feed (G:F). Blood was collected from all steers on days 0, 28, 56, 84, and 126. Liver samples were collected on days −3 or −2 and day 123 or 124. Data were analyzed using ProcMixed of SAS (experimental unit = steer; fixed effect = treatment and block). Plasma Cu was analyzed as repeated measures (repeated effect = day). Plasma and liver Cu concentrations were regressed against total Cu intake using ProcGLM to calculate relative bioavailability of GLY. Final BW and overall ADG were greatest for CS5 and CS10 and least for CON and GLY5 (P = 0.01). Overall, DMI was not affected by treatment (P = 0.14), but overall G:F tended to be greatest for CS5, CS10, and GLY5 and least for CON (P = 0.08). Total and supplemental Cu intake was greatest for steers supplemented either source at 10 mg Cu/kg DM and least for CON (P < 0.01). However, total and supplemental Cu intake was greater for CS5 than GLY5 (P < 0.01). Final liver Cu concentrations were greatest for CS10, least for CON, CS5, and CS10, and intermediate for GLY10 (P < 0.01). Final plasma Cu was greatest for steers supplemented either source at 10 mg Cu/kg DM (P < 0.01). Relative bioavailability of GLY was 82% compared to CS based on liver Cu (P < 0.01) but did not differ based on plasma Cu (P = 0.60). The lesser bioavailability of GLY relative to CS could be due to a high concentration of dietary antagonists and lower solubility of GLY (68.9% relative to CS) in pH conditions (5.2) similar to the ruminal pH of a beef animal consuming a high concentrate diet. Future studies should examine the effects of bis-glycinate bound Cu fed in blended combination with inorganic Cu sulfate to determine the most effective blend of sources for feedlot cattle experiencing varying amounts of dietary Cu antagonists.