Stomatal frequency (SF) of five date palm (Phoenix dactylifera L.) cultivars was determined under three different treatments: in vitro (control), in vitro polyethylene glycol (PEG) treated for one week (acclimatized), and greenhouse-grown plants. The SF of greenhouse leaves (94.46 stomata mm-2) was significantly higher than the control plantlets (58.48). The PEG treatment did not increase the number of stomata mm-2 of in vitro plantlets. Four hours after detachment, leaves of greenhouse-grown plants had 98% of the stomata closed while only 20% of the stomata of in vitro leaves were closed. The PEG treatment did not affect the stomata] closure mechanism of in vitro plantlets. Photomicrographs of leaf imprints showed no differences in stomatal size and surface topography among the three treatments. Excessive water loss from leaves of transplanted in vitro plantlets may be due mainly to stomata which remain open.