Comparisons of quantitative approaches for assessing microglial morphology reveal inconsistencies, ecological fallacy, and a need for standardization

Microglia morphology is used as a measure of neuroinflammation and pathology, but different methods to quantify microglia morphology are frequently employed across neuroscience. For reliable inference, it is critical that microglial morphology is accurately quantified and that results can be easily interpreted and compared across studies. We applied five of the most commonly used ImageJ-based methods for quantifying the microglial morphological response to a stimulus to identical photomicrographs and isolated microglial cells, which allowed for direct comparisons of the specificity and reliability of each method.