Skeletal muscle form and function has fascinated scientists for centuries. Our understanding of muscle function has long been driven by advancements in imaging techniques. For example, the sliding filament theory of muscle, which is now widely leveraged in biomechanics research, stemmed from observations made possible by scanning electron microscopy. Over the last 50 years, advancing in medical imaging, combined with ingenuity and creativity of biomechanists, have provide a wealth of new and important insights into in vivo human muscle function. Incorporation of in vivo imaging has also advanced computational modeling and allowed our research to have an impact in many clinical populations. While this review does not provide a comprehensive or meta-analysis of the all the in vivo muscle imaging work over the last five decades, it provides a narrative about the past, present, and future of in vivo muscle imaging.