Objectives: Investigate how prosthesis of different lengths affect tympanic membrane (TM) and footplate motion in a temporal bone model of tympanoplasty with Laser-Doppler vibrometry (LDV) and opto-electronic holography (OEH) measurements. Methods: Four human temporal bones without history of otologic disease were prepared using universal precautions. Preparation included removal of the bony external auditory canal to expose the majority of the TM surface, and canal wall-up mastoidectomy with wide posterior tympanotomy. Retroreflective balls were placed on the posterior crus of the stapes. Control measurements were made in bones with intact ossicular chain, then the incus and the stapes suprastructure were removed with a KTP laser. The ossicular chain was reconstructed using a total ossicular replacement prosthesis (TORP). LDV measurements of the footplate motions, together with OEH measurements of TM surface and umbo motions, were made before and after placement of different length TORPs. The effect of different sizes of a cartilage disk interposed between the TM and the prosthesis was also investigated. Results: A ‘Best Fit’ TORP with a small cartilage disk gave the highest stapes velocities. Either TORP with a small disk produced larger OEH determined displacements of the whole TM surface, particularly at low frequency. In general, a smaller cartilage disk resulted in higher mobility of the TM and stapes. Conclusions: Preliminary results indicate reconstruction length and the size of the cartilaginous disk differentially affect the mobility of the TM and the stapes. However, there is significant variation in the acoustic response of similarly reconstructed ears, even in this relatively well-controlled preparation.