Introduction: A number of studies defined normal reference values of three-dimensional (3D) speckle-tracking echocardiography (3DSTE)-derived left ventricular (LV) strains. The present study aimed to quantify normal reference values of LV strains in healthy adult population in real clinical world settings in different age groups, and to determine ageand gender-dependence of these variables in a high volume single center. Methods: The present prospective study included 296 healthy adult subjects. Among them, 124 were excluded due to inferior image quality during the 6-year recruitment period (2011-2017). The remaining population was further divided into four subgroups based on age decades. Results: While global radial (RS) strain showed an increase-decrease-increase pattern with age, circumferential, longitudinal, and area strains (CS, LS, AS, respectively) were nonsignificantly lower in older ages. Only global LV-LS showed gender-dependency with higher values in females. Although moderately higher RS and 3DS and lower LS, and AS were observed in males, clear gender-dependency could not be detected in different age decades. Conclusions: This study provides normal reference values of 3DSTE-derived global, segmental, mean segmental, and regional LV strains in healthy adult subjects based on real-life clinical experience. Age-, gender-, and functional nonuniformity of LV strains were also defined.