The "neutral guest" protocol described above (31 lambda windows) was used to run all simulations in the cyclodextrin dataset with guest g1, for 16 ns per lambda window when free energy estimates appeared converged. On the other hand, the "charged guest" protocol (61 lambda windows) was used for the remaining host-guest systems across all datasets since all other guests bore a formal charge. In this case, simulations were run until free energy estimates apparently converged or up to 30 ns per lambda window, which ever came first. First, to determine feasible cross application of the "charged guest" protocol to different systems (GDCC and TrimerTrip datasets), the charged protocol was tested on OA-g2 and clip-g11. Experimental data for OA-g2 was available from a previous SAMPL challenge, so this was an ideal system to test the protocol. The OA-g2 test resulted in predicted free energy within 4 kcal/mol, after running the simulation to 26 ns per window. A health report for the OA-g2 simulation showed reasonable mixing between replicas, and there was apparent convergence. However sampling of replicas in individual states was not ideal. For the clip-g11 test simulation (for TrimerTrip dataset), the protocol was initially deemed reasonable based on YANK's health report (with mixing_cutoff and mixing_warning_threshold options at default 0.05 and 0.9 settings, respectively) which can detect insufficient replica mixing or number of swaps between states and thus issue warnings.