Vitrification is a promising alternative to tissue preservation, which will greatly avoid the ice-crystal formation and circumvent the hazardous effects associated with ice formation during the entire procedures. In this study, we evaluate the effect of vitreous cryopreservation of rabbit trachea by comparing the vitrification procedure with the conventional computer-programmed slow freezing approaches. Harvested tracheae were tailored and divided into groups and cryopreserved by vitrification and programmed freezing, respectively. The morphology and ultrastructure of the thawed tracheal fragments were examined by using HE staining, TUNEL test, transmission electron microscopy (TEM) and scanning electron microscopy (SEM) to assess the integrity of the tracheal fragments. The morphological studies demonstrated both cryopreservation procedure retain the interity of trachea, and that epithelium mucosae, cilia and cartilage cells were in good shape. Compared with slow freezing methods, vitrification was less detrimental to cartilage cells and had a higher survival rate of chondrocytes and coverage of epithelium and cilia. Therefore, vitrification method is a more satisfactory method to preserve trachea, the survival of chondrocytes in situ in cartilage tissue is adequate, and respiratory epithelium is soundly preserved.