Two-dimensional (2D) layered materials are studied in efforts to discover new compounds and for their fascinating properties engendered by their sheet-like structure and tunable surfaces. MXenes are an emergent class of layered, synthesized transition metal carbides and carbonitrides that are useful in addressing the formidable challenges of sensing at the energy-water nexus. This work reports systematic structural and electrochemical properties of titanium carbide (Ti3C2Tx) MXenes revealed by varying interlayer spacing, flake thickness and lateral size under different electrolytes. In addition to traditional electrode kinetics, we utilized surface sensitive scanning electrochemical microscopy (SECM) to gain a more complete understanding of rich MXene surface chemistry and corresponding knowledge about the physicochemical processes, including inherent electrochemistry and heterogeneous charge transfer characteristics, at electrode/electrolyte (solid/liquid) interfaces. We employed electron microscopy, ultraviolet–visible (UV–Vis) absorption spectroscopy, x-ray diffraction, and Raman spectroscopy to determine surface morphology, microscopic and electronic structure and lattice vibrational properties. It is shown that Ti3C2Tx or, specifically, transition metal Ti, undergoes irreversible oxidation and lithiation in a positive potential window, which strongly depends on the flake thickness and type (aqueous versus organic) of the electrolyte. Multi-layered and smaller Ti3C2Tx flakes exhibit faster electron transfer kinetics (kET = 1.2 cm s−1) with a potassium ferrocyanide [Fe(CN)6]4−/3− redox probe, compared to few-layered Ti3C2Tx (kET = 0.3 cm s−1) in aqueous and organic electrolyte (kET = 4.9 cm s−1) with a [Fe(CN)6]4−/3− redox probe, and compared to a few-layered Ti3C2Tx (kET = 0.9 cm s−1). In addition, the few-layered free standing Ti3C2Tx film electrode remains intact following irreversible oxidation. These properties help to establish structure–property-electroactivity relationships among different types of Ti3C2Tx MXenes.