The field of mass spectrometry imaging (MSI) is constantly evolving to analyze a diverse array of biological systems. A common goal is the need to resolve cellular and subcellular heterogeneity with high spatial resolution. As the field continues to progress towards high spatial resolution, other parameters must be considered when developing a practical method. Here, we discuss the impacts of high spatial resolution on the time of acquisition and the associated implications they have on an MSI analysis (e.g., area of the region of interest). This work presents a brief tutorial serving to evaluate high spatial resolution MSI relative to time of acquisition and data file size.