Herceptin (Trastuzumab) is a widely used and effective drug for the treatment of Her2+ breast cancer but its cardiotoxic side effects require regular monitoring by echocardiography. A 10% reduction in left ventricular ejection fraction can lead to suspension of treatment and therefore has significant implications for patient prognosis in terms of cardiac and cancer outcomes. Assessment of LV function by conventional 2D biplane method of discs (2DEF) has limitations in accuracy and reproducibility. Global longitudinal strain (GLS) is becoming more widely available and user friendly. It has been shown to demonstrate myocardial damage earlier in treatment than 2DEF, allowing the option of pharmacological intervention at a pre-clinical stage and preventing the interruption of Herceptin. This study compares the reproducibility of GLS with that of 2DEF in a routine clinical environment. Fifty echocardiograms performed on female patients undergoing Herceptin treatment were used to measure both 2DEF and GLS within the recommended standard appointment time of 40 min. The data were re-measured (blind) by the same operator a minimum of 14 days later to determine intra-operator variation. These data were also measured by a second operator (blind), to assess inter-operator variation. Analysis by direct comparison, intra-class correlation (ICC), coefficient of variation (CV) and Bland–Altman plots demonstrated that GLS is a more reproducible measurement than 2DEF. This is important to prevent clinical decisions being erroneously based on variation in operator measurement. The investigation also shows that with advances in machine software this is a practical addition to routine assessment rather than merely a research tool.