Abstract During bias-enhanced nucleation (BEN) of diamond on iridium, an ultrathin carbon nucleation layer is deposited. The nature of the nucleation centers or nuclei which gather in discrete islands is investigated in the present work by subjecting the as-biased samples to different treatments. In high vacuum annealing experiments, the structures proved completely stable up to 1000 °C. Above this temperature, the nuclei were destroyed and the iridium was etched within the domain areas. Low energy oxygen and xenon ion beams were then used to etch very shallow craters of varying depth. The induced modifications of the surface and the influence on the nucleation density can be understood on the basis of calculations which simulate the damage caused by the ion bombardment. Finally, the stability of the nucleation structures under hot chromo sulfuric acid treatment was compared with the corresponding behavior of a thick amorphous carbon layer. In all the experiments, the nucleation structures formed during BEN behaved similar to diamond situated at the surface of the iridium buffer layer possibly covered by a thin stable amorphous carbon layer.