Experimentally measured assembly indices are required to determine the threshold for life.

Assembly theory (AT) aims to distinguish living from non-living systems by explaining and quantifying selection and evolution. The theory proposes that the degree of assembly depends on the number of complex objects, with complexity measured using a combination of the object's assembly index (AI) and its abundance. We previously provided experimental evidence supporting AT's predictive power, finding that abiotic systems do not randomly produce organic molecules with an AI greater than approximately 15 in detectable amounts. Hazen et al. (Hazen et al. 2024 J. R. Soc. Interface 21, 20230632. (doi:10.1098/rsif.2023.0632)) proposed inorganic molecules that theoretically have AIs greater than 15, suggesting similar complexity to biological molecules. However, our AIs are experimentally measured for organic, covalently bonded molecules, whereas Hazen's are theoretical, derived from crystal structures of charged units that are not isolable in solution. This distinction underscores the challenge in experimentally validating theoretical AIs.