Purpose: The purpose of this study was to examine the effect of the 226-Hz probe level on the acoustic stapedius reflex threshold. Method: Contralateral reflex thresholds for a 1000-Hz pure-tone stimulus were obtained from 40 young adults with normal hearing using an experimental system at four 226-Hz probe levels (70, 75, 80, and 85 dB SPL) with a repeated measures experimental design. Results: A repeated measures analysis of variance revealed that as the probe level increased over this 15-dB range, there was a mean decrease in reflex threshold of 2.5 dB SPL (p less than 0.001), with the largest decrease for an individual participant of 12 dB. Conclusion: The results suggest that the level of the 226-Hz probe tone as used in clinical reflex measurements and as specified in the American National Standards Institute (1987) standard is sufficiently high to affect the acoustic reflex by facilitating its activation. The effect is small, on average, but may be of clinical significance for some individuals.