In the search for possible causes of unfavorable results after stapes surgery, the study reported here focused on the anterior mallear ligament, since it has been previously reported that partial mallear fixation (PMF) leads to functional failure in 38% of cases of stapes revision surgery. The aims of the study were to identify effective methods for the diagnosis of PMF and experimentally assess the conductive hearing loss that results from PMF. The study included vibration amplitude measurements of the ossicles by laser Doppler interferometry (LDI) in 19 patients and 5 fresh human temporal bone (TB) specimens. Analysis of their dynamic behavior was performed by finite element modeling (FEM). Similar, significant changes of manubrium vibration patterns for PMF were found by FEM calculations, in TB experiments, and in patients. We could identify PMF either before operation, using LDI, or during operation, by manual palpation. In the TB experiments and FEM calculations, the attenuation of the stapes displacement due to an isolated PMF was approximately 10 dB and frequency-dependent. Untreated anterior mallear ligament fixation produced a persistent air-bone gap of approximately 10 dB after stapedioplasty.