Objective To establish a new method for quantitative observation of inner ear hair cells, neurons and nerve fibers from sagittal sections of temporal bones. Methods Temporal bones of four CBA/CaJ mice at 45 days of age were removed under deep anesthesia. The inner ear cavity was perfused with 2.5% glutaraldehyde and then immersed in the fixative for 6 hours. The temporal bones were rinsed with PBS, and immersed in 2% osmium tetroxide for 2 hours. After routine decalcification, dehydration, and epoxy resin embedding, semi-thin sections (2 μm) were prepared along the sagittal plane of the temporal bones and stained with 0.5% toluidine blue. Quantitative observations of hair cells, neurons and nerve fibers were performed in a small field of view under an optical microscope. Results Collection and observations began with planar sections parallel to the medial wall of the temporal bone. Upon sectioning through the medial wall to access the cochlear nerve canal and superior vestibular nerve canal, cross-sections of the central nerve fibers of the corresponding cochlear nerve bundles and inferior vestibular nerve bundles were obtained. Sections through the macula of the utricle, the crista ampullae of the superior and lateral semicircular canals revealed cross-sectional profiles of hair cells in these structures, as well as the soma of superior vestibular neurons and their peripheral and central nerve fibers. Similarly, sections through the macula of the saccule and the crista ampulla of the posterior semicircular canal were also profiled cross-sectional views of their hair cells, along with the soma of inferior vestibular neurons and their peripheral and central nerve fibers. Cross-sections of the bony spiral lamina of the cochlea exposed the peripheral nerve fibers of the spiral ganglion, while sections through the center axis of modiolus revealed the cross-section of cochlear hair cells and the soma of spiral ganglion neurons. Quantitative analysis demonstrated that the average density of hair cells in vestibular sensory epithelium of normal mice was (18.67±1.88)/180 μm. The number of peripheral nerve fibers within a single habenula perforata in the middle turn of cochlea was 57.83±9.09. The average density of spiral ganglion neurons of the cochlea, superior and inferior vestibular ganglion neurons was (20.39±2.23)/0.007 5 mm². The average density of central nerve fibers in the cochlear nerve, inferior and superior vestibular nerve was (497.06±25.28)/0.007 5 mm². Conclusion The sagittal plane of the temporal bone provides a favorable perspective for comprehensive cytological and quantitative assessments of the inner ear nervous system.