Echocardiographic assessment of Left ventricular systolic function is traditionally being performed by estimation of fractional shortening (FS) and ejection fraction (EF). Speckle tracking echocardiography (STE) is a promising tool for assessment of myocardial function. The aim of this study is to evaluate the global longitudinal strain (GLS) using 2D-STE in healthy neonates to establish normal reference ranges. It is a retrospective study through an analysis of transthoracic echocardiogram of normal healthy neonates. We enrolled all neonates in our institution from January 1st, 2021 to February 28th, 2021. 2-D STE was used to assess left ventricular GLS from the apical views. 185 neonates were enrolled. Mean value for left ventricle GLS (%) was -19.9±1.2, GLS-derived EF (%) was 60.0±2.7; while the left ventricle EF by biplane Simpson's method (%) was 61.0±3. There is a good positive correlation between the Left Ventricle EF by biplane Simpson's method and EF by 2-D STE, which was statistically significant (r=.294,n=102,p=.003). Apical 4-chamber longitudinal strain and strain-derived EF is significantly correlated with GLS and bi-plan EF respectively. 2-STE is feasible technique for analyzing newborn myocardial systolic function. The normal range of GLS in neonates is not much different than reported for the pediatric. There is a good positive correlation between the Left Ventricle EF by 2-D STE and EF by biplane method.