We thoroughly characterized a system for thermal separation of organic carbon (OC) and elemental carbon (EC) for subsequent radiocarbon analysis. Different organic compounds as well as ambient aerosol filter samples were introduced into an oven system and combusted to CO2 in pure O2. The main objective was to test which combustion times and temperatures are best suited to separate OC and EC. The final separation step for OC was combustion at 360 °C for 15 min. Combustion at this temperature proved enough to remove several organic test substances from the filter (including high molecular weight humic acid) but did not remove substantial amounts of EC. For isolation of EC, OC first needs to be completely removed from the filter. This was achieved by water extraction of the filter, followed by combustion of the water insoluble OC at 360 °C and combustion at an intermediate temperature step of 2 min at 450 °C. This last step removed the most refractory OC together with some EC. Finally, the remaining EC was combusted to CO2 at 650 °C. The recovery of black carbon after the intermediate 450 °C step was approximately 80%. Several tests provided strong evidence that OC was removed efficiently during the intermediate temperature step: (i) brown carbon, indicative of refractory OC, was removed; (ii) the fraction modern of EC did not decrease significantly if the temperature of the intermediate step was further increased. Based on tests with various organic compounds, we estimated that charred organic carbon could contribute 4–8% to an elemental carbon sample that was isolated according to our method.