The commonly used methods to quantify axon numbers and mean area include manual and semiautomated procedures. The authors introduce a new fully automated method of morphometric analysis using ImageJ and Paint.net software to improve efficiency and accuracy. A total of six rat sciatic nerves were examined for their axon numbers and mean axon area by comparing the manual method or semiautomated MetaVue method with the new ImageJ method. It was observed that the number of axons for manual counting and ImageJ were 4,630 ± 403 and 4,779 ± 352, respectively, and the difference was not statistically significant (p > 0.5, t-test). The mean axon area measured was 13.44 ± 2.62 µm2 for MetaVue and 8.87 ± 0.78 µm2 for ImageJ, respectively, and the difference was statistically significant (p < 0.01, t-test). The standard error and coefficient of variation of MetaVue were 1.07 and 0.195; and for ImageJ were 0.32 and 0.087. The authors conclude that their new approach demonstrates improved convenience, time efficiency, accuracy, and less operator error or bias.
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