Purpose: Ultrasonography has been extensively explored in dentistry because of its several diagnostic advantages. The purpose of this study was to determine the thickness of the maxillary attached gingiva with the use of a high-frequency ultrasound in subjects with sex-, age- and smoking-related differences. Materials and Methods: A total of 100 healthy subjects (70 women and 30 men, age range: 18 to 55 years) were included. A special B-scan unit was used to measure the soft tissue thickness in the buccal and palatal attached gingiva in the maxillary arch. Aquasonic 100 Ultrasound Gel (Parker Laboratories) was used as coupling medium. Student t test was used for statistical analysis of the subgroups with different parameters with significance set at P .05. Results: A total of 2,734 measurements showed that male nonsmokers have a significantly thicker fixed gingiva than nonsmoking women and that age does not seem to have great influence on the thickness of the gingiva. Smoking had a significant effect, but only on the oral maxillary gingival tissues of women. The palatal gingiva was found to be significantly thicker in female smokers than in nonsmoking females. Conclusions: Measurement of gingival thickness for different purposes using a B-scan ultrasonic device appears to be a reliable method. Sex- and smoking-related differences in the gingival thickness exist only on the palatal side among women.