Assessing fractional hair cell survival in archival human temporal bones

Objectives/Hypothesis Histopathological analysis of hair cell survival in human temporal bone sections has historically been binarized such that each hair cell row is rated as either present or absent, thereby greatly underestimating the amount of hair cell loss. Here, we describe and validate a technique to reliably assess fractional hair cell survival in archival sections stained with hematoxylin and eosin (H&E) using high‐resolution light microscopy and optical sectioning. Study Design Technique validation. Methods Hair cell counts in archival temporal bone slide sets were performed by several observers using either differential interference contrast (DIC) or confocal microscopy of the endogenous eosin fluorescence in hair cells. As a further cross‐check, additional decelloidinized sections were immunostained with hair cell markers myosin VI and VIIa. Results Cuticular plates and stereocilia bundles are routinely resolvable in DIC imaging of archival H&E‐stained human material using standard research‐grade microscopes, allowing highly accurate counts of fractional hair cell survival that are reproducible across observer and can be verified by confocal microscopy. Conclusions Reanalysis of cases from the classic temporal bone literature on presbycusis suggests that, contrary to prior reports, differences in audiometric patterns may be well explained by the patterns of hair cell loss. Level of Evidence NA Laryngoscope , 130:487–495, 2020