Purpose: The authors investigated the reliability of myotonometry-measured muscle tone in the thenar and perineal muscles. Methods: Participants were women aged 18-50 years who were asymptomatic for thumb and pelvic floor dysfunction (interrater study n=20; intrarater study n=43) or who were symptomatic for vulvodynia (interrater study n=14; intrarater study n=32). Mechanical properties (stiffness, frequency, decrement, relaxation time, and creep) of the muscles were measured using a myotonometer (MyotonPRO) while the muscles were in a relaxed state. Measures were performed twice by two assessors. Intra- and interrater reliability were determined using intra-class correlation coefficients (ICCs) and absolute reliability using the standard error of measurement and a minimum detectable change. Results: The primary property of interest, muscle stiffness, showed very good interrater (ICC 0.85-0.86) and intrarater (ICC 0.82-0.88) reliability in the thenar eminence. In the perineal muscles, reliability results ranged from good to very good for interrater (ICC 0.70-0.86) and intrarater (ICC 0.80-0.91) reliability for muscle stiffness. Absolute reliability was confirmed, with all measures showing minimal variance. Conclusions: Muscle stiffness of the smaller muscles of the body can be reliably measured using the MyotonPRO. The device could be used as a reference standard in the development of a digital palpation scale that would facilitate accurate diagnosis of muscle tone.