The lack of a polymer electrolyte membrane (PEM, e.g. Nafion) in membraneless, laminar flow-based micro fuel cells (LF-FCs) eliminates several PEM-related issues such as fuel crossover, cathode flooding, and anode dry-out, as we reported previously. This paper explores the media flexibility of LF-FCs by working in acidic and alkaline media, as well under “mixed-media” conditions in which the anode is in acidic media while the cathode is in alkali, or vice versa. Operating a fuel cell under alkaline conditions has positive effects on the reaction kinetics, both at the anode and cathode, while the cell performance under “mixed-media” conditions offers an opportunity to increase the maximum achievable open cell potential (OCP). The lack of media-related constraints and the simplicity of the LF-FC design allow for these experiments to be performed consecutively in a single LF-FC without changing the system, except for altering the composition/pH of the fuel and oxidant stream. The performance of LF-FCs operated with different media is described and compared.