The hearing organ includes the outer, middle, and inner ear. The middle ear transfers sounds into the inner. Wideband energy reflectance (ER) is a new clinical procedure that assesses middle ear function. It gives information for a broad frequency range, an advantage over currently used 226 Hz tympanometry procedure. Previous studies reported that repetitive testing of single-frequency tympanometry altered outcomes. We investigated ER measures with eight consecutive wideband tympanometry tests in normal adult ears. This study has provided the first set of data showing significant frequency-specific change in ER measures. For further research that requires repeated measurements, this effect needs to be minimized to improve reliability.