Chemical gradients drive a diverse set of biological processes ranging from nerve transduction to ovulation. At present, the most common method for quantifying chemical gradients is microscopy. Here, a new concept for probing spatial and temporal chemical gradients is reported that uses a multi-layer microfluidic device to measure analyte concentration as a function of lateral position in a microfluidic channel using electrochemistry in a format that is readily adaptable to multi-analyte sensing.