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Dianthus caryophyllus cv. Nelken was cultured in vitro under different ventilation rates (0.11, 0.21, 0.68 and 0.86 changes h⁻¹). Ventilation modified the anatomical characteristics of shoots and leaves described for plants grown in non-ventilated vessels: the cuticle became thicker, there was a decreased cell size and intracellular space size. Also, there were more photosynthetic and supportive tissues, including thicker cell walls. Increased ventilation promoted in vitro hardening of micropropagated carnation shoots, and pushed the culture-induced phenotype closer to that of ex vitro acclimatized plants. Anatomical variability of in vitro-grown plants was demonstrated to be a consequence of ventilation.