The timing, causes and extent of mantle heterogeneity preserved in the ∼492 Ma Shetland Ophiolite Complex (Scotland) are evaluated using Re-Os isotope and whole rock highly siderophile element (HSE: Os, Ir, Ru, Pt, Pd, Re) abundance measurements of a suite of eight chromitites and 21 serpentinised harzburgites and dunites. Shetland dunites have more variable initial 187Os/188Os, as well as absolute and relative abundances of the HSE, compared to spatially associated harzburgites. As is common for ophiolitic peridotites, the harzburgites (γOs492Ma of −5.3 to +2.6) preserve evidence for a Mesoproterozoic depletion event, but are dominated by contemporary chondritic, ambient upper mantle compositions. The dunites have γOs492Ma values ranging between −3.3 and +12.4, reflecting dunite formation by higher degrees of melt interaction with mantle rock than for the spatially associated harzburgites.