Each system consisted of two production runs, starting from both the uncompressed and buckled bilayers. The uncompressed bilayer production run was under anisotropic conditions, with compressibility set to 0 in the y-dimension such that the bilayer length can only fluctuate in the x-dimension. The buckled bilayer production run was under semi-isotropic conditions, with compressibility in and x y set to 0, fixing the buckle in place. The bending modulus can be calculated from the force exerted by the buckled membrane in the x-dimension that is proportional to kc according to: