Standardization of postmortem human brainstem along the rostrocaudal axis to accommodate for heterogeneity in samples.

Human postmortem brain tissues provide an indispensable resource that is crucial for the understanding of neurological conditions, whether related to pathology subtype, burden, distribution or cell-type specificity. Pathology staging protocols provide guidelines for standardized sampling of brain tissues, but cover only a subset of regions affected by pathologies. Thus, to study how various neuropathologies and cell types in highly specialized circuit nodes correlate with functions specifically served by these nodes, additional protocols are necessary. This especially applies to brainstem tissues due to the small dimension of regions of interest and interindividual variability of specimens, whether due to procurement or intrinsic differences. Here we systematically assessed factors contributing to heterogeneity in the length of whole brainstem samples and then presented a standardized approach to reproducibly assign rostrocaudal levels, with standardization relying upon readily identifiable internal landmarks. We validated this approach using postmortem MRI imaging. Standardized brainstem length correlated positively with subject height and negatively with subject age of death. By providing a reference series, reproducible levels can be assigned to individual histological sections or MRI images, i.e. when full brainstem specimens are not available and irrespective of platform, promoting reproducibility.