Heart failure (HF) is a major global health problem that affects millions of people every year. HF can result from various cardiomyopathies that result in impaired contractile function of cardiomyocytes (CMs) and cardiac sarcomere remodeling. Specifically, the cardiac sarcomere consists of thick and thin filaments that slide past each other to generate force in response to hemodynamic stimuli. Sarcomere remodeling is a process that adapts the sarcomere length and number to changing conditions, driving both physiological and pathological changes. However, the mechanisms and consequences of sarcomere remodeling are not yet fully understood, especially in humans. While animal models have limitations in terms of human relevance and imaging depth, the in vitro models made of myocardium-isolated or stem cell-derived CMs can offer more control and specificity to help us better decipher the mechanism of sarcomere remodeling-associated cardiomyopathies. Moreover, sarcomere remodeling has been studied by various live-cell and live-sarcomere visualization modalities to correlate the sarcomere dynamics to a variety of applications, such as the unique pathogenesis of cardiomyopathies, the maturation process, and validation of engineered cardiac constructs. In this chapter, we introduced the current state-of-the-art sarcomere imaging techniques, focusing on the advantages and challenges of imaging live CMs in non-invasive ways for studying sarcomere remodeling in various in vitro culture models. We further highlighted the applications of sarcomere remodeling in disease modeling of cardiac diseases and developing novel therapeutics. Last, we discussed the current limitations and future perspectives of studying sarcomere remodeling with more advancements in non-obstructive live imaging technologies.