The mechanism by which syntectonic hydrothermal veins form is widely debated, with some workers suggesting that certain vein textures are related to specific fluid flow regimes. Central to the debate is whether vein formation involves advective fluid flow, or occurs by local diffusion of material from the surrounding wall rock. To address this issue, we integrated textural observations and microchemical analyses of a hydrothermal vein from the Lachlan Orogen, southeast Australia, to reveal information about vein growth history, changes in fluid chemistry, and the evolution of fluid flow pathways during vein growth. The study area is part of a regional-scale fault-fracture network in an interbedded limestone-shale sequence, which formed at depths of ∼ 5-10 km (∼ 150-200 °C) during late Devonian crustal shortening. This integrated approach demonstrates that the zonation of textures, Sr isotopes, stable isotopes (C, O), and trace and rare earth elements is distinctly asymmetrical about the median growth-line of the vein. δ18O values in vein calcite (17.0-18.8‰, VSMOW) are lower than those in surrounding unaltered limestones (23-25‰, VSMOW), and vary systematically across the vein. In contrast, δ13C values are relatively constant across most of the vein, but become markedly depleted (ca. 4‰) immediately adjacent to the wall rock. This strong depletion in δ13C was probably caused by the influx of more oxidised fluids during the latest stages of vein growth. Strontium isotope ratios (87Sr/86Sr) vary between 0.70912 and 0.70931 across the vein. Abrubt changes in 87Sr/86Sr, δ18O, Ce/Ce⁎, Eu/Eu⁎ and trace element concentrations indicate that vein growth was accompanied by stepwise changes in the fluid flow pathway and consequent variations in fluid chemistry. Taken together, our findings are not consistent with growth of fibrous antitaxial veins by diffusional transfer of material from the surrounding wall rock. Instead, they suggest that externally sourced fluids migrated along episodically changing fracture-controlled flow pathways. This has implications for the dynamics of crustal permeability and mineralisation.