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By conducting a systematic study of model lipid membranes using the atomic force microscopy (AFM) indentation, we demonstrate the importance of an experimental protocol on the determination of their mechanical parameters. We refine the experimental approach by analyzing the influence of the contact mechanics models used to process the data, substrate preparation, and indenter geometry. We show that both bending rigidity and area compressibility can be determined from a single AFM indentation measurement.The extent of intersystem crossin