Myosin VIIa Monoclonal Antibody Labels Normal and Dying Hair Cells in the Gentamicin Treated Avian Cochlea

Objectives: Myosin VIIa is a motor protein critical to the survival of cochlear hair cells. Mutations in the myosin VIIa gene cause hair cell death and deafness. We will determine if a monoclonal antibody to myosin VIIa labels normal and dying hair cells similarly to a well characterized polyclonal myosin VIIa antibody used routinely for identifying hair cells. Study Design: Two week old chicks get a single gentamicin injection to induce hair cell loss in the basal 30% of the cochlea. Birds survived for 1-4 days after the gentamicin when their cochleae were extracted and processed for immunocytochemical analysis. Methods: Cochleae were labeled with either a monoclonal or polyclonal primary antibody to myosin VIIa followed by an appropriate secondary antibody. The cochleae were counterstained with phalloidin, a fluorescently tagged probe for actin filaments. The whole mount cochleae were examined with a Zeiss 510 Meta confocal microscope. Results: The monoclonal myosin VIIa antibody labeled normal hair cells in a manner identical to the polyclonal antibody. It also labeled myosin reorganization in dying hair cells being ejected from the cochlea and afterwards when they were dead but trapped in the overlying tectorial membrane. Conclusions: Both the polyclonal and monoclonal antibodies we tested equivalently and specifically label normal and dying hair cells in the chick cochlea. This enables much more flexibility in co-labeling hair cells with the monoclonal myosin VIIa antibody and other polyclonal antibodies to a wide variety of hair cell structural, developmental and damage induced proteins.