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Cardiomyocytes (CMs), the cells responsible for the contraction of the heart, experience mechanical forces such as active and resting tension as the cell contracts and relaxes. Active tension is generated by the contractile cytoskeleton, which is composed of actin-myosin arrays called sarcomeres. Resting tension is the tension that results when spring-like elements are stretched beyond their resting length, and this resting tension determines how forceful a contraction will be. The ability to investigate how resting tension is regulated has been a challenge due to the lack of a robust method to measure resting tension at the cell level. Previous methods involve stretching an individual cell and measuring the resulting resting tension; however, this method is low throughput and invasive. Single-cell Traction Force Microscopy (TFM) serves as a promising alternative method to measure resting tension due to the ability to …