Neottopteris nidus prothalli and green globular bodies (GGBs) were successfully cryopreserved by droplet-vitrification. Prothalli were subjected to different treatments. The following parameters were studied: the age of in vitro mother plants from which prothalli were originated (30 to 90-day old), length of exposure to loading solution (LS) (20 to 40 min) and length of exposure to the plant vitrification solution (PVS2) (10 to 55 min). N. nidus GGBs and GGBs in suspension were subjected to PVS2 for 20, 30 and 40 min before liquid nitrogen exposure. The highest prothalli regrowth (92%) occurred when they were exposed for 40 min to LS, followed by 20 min to PVS2 and when they originated from non-preconditioned 45-day old mother plants. The highest GGB (100%) and GGB suspension regrowth (100%) after cryopreservation occurred when they were exposed to PVS2 for either 20 min or 40 min.