How a cell senses, responds, and moves toward, or away from an external cue is central to many biological and medical phenomena including morphogenesis, immune response, and cancer metastasis. Many eukaryotic cells have internal sensory mechanisms that allow them to sense these cues, often in the form of gradients of chemoattractant, voltage, or mechanical stress, and bias their motion in a specific direction. In this study, a new method for using microfluidics to study the electrotactic migration of cells is presented. Electrotaxis (also known as galvanotaxis) is the phenomenon by which cells bias their motion directionally in response to an externally applied electrical field. In this work, we present a new flow-based, salt bridge-free microfluidic device for imaging and quantifying cell motility and intracellular ion activity during electrotaxis. To eliminate salt bridges, we used a low nanoliter flow rate to slowly drive Faradaic waste products away from and out of the electrotaxis zone. This cell migration zone consisted of an array of fluidic confinement channels approximately 2 μm in thickness. This confined height served to insulate the migrating cells from the electric field at the top and bottom of the cell, such that only the two-dimensional perimeter of the cells interacted with the electrical source. We demonstrate the ability to quantify the electrotactic velocity of migrating Dictyostelium discoideum cells and show how this confined design facilitates the imaging and quantification of the ion activity of electrotaxing cells. Finally, by spatially imaging the calcium concentration within these cells, we demonstrate that intracellular calcium preferentially translocates to the leading edge of migrating Dictyostelium cells during electrotaxis but does not exhibit this behavior during migration by chemotaxis in a gradient of cyclic adenosine 3′,5′-monophosphate or when cells freely migrate in the absence of an external cue.