Despite the imperfect agreement between the fitted 𝐾 𝐷 and 𝛼 parameters to the phosphate series experimental data manifest by Cu II precipitating from solution, bivariate fitting of repeat measurements (see ESI) indicated that the positive cooperativity and reduced initial binding affinity compared to the Tris-HCl buffer series was reproducible. Additionally, scaling experimental Cu II concentration by 0.65 for the phosphate and 0.85 for the Tris-HCl buffer series yielded global RMSD minima upon reprocessing (see ESI). This observation is consistent with the CW-EPR data (see ESI) for the phosphate buffer series that only ~60% of Cu II is retained in the solution. While for the Tris-HCl buffer series, scaling by a factor of 0.85 corresponds to a shift in optimised dimer formation from a metal:protein ratio of 3:5 to 1:2, as expected for negative cooperativity.