Prior to network generation, λ allocations were benchmarked in the context of RBFE statistical fluctuation (figure ) in the solvated phase for six highly reliable and six highly unreliable perturbations in RBFE-Space. The statistical fluctuation (here expressed as SEM ∆G solvated of 5 replicates) was recorded at increasing numbers of equidistant λ windows used for MBAR analysis: 3, 5, 9, 17 and 33. For both types of perturbations an exponential decay in SEM ∆G solvated was observed; typically convergence was reached at 15-20 λ windows, suggesting further sampling is likely not necessary in RBFE calculations with SOMD for the solvated phase, even for highly unreliable perturbations.