Comparison of two methods for linear measurement of ossified tissue in cleared mouse fetuses

Two methods of measuring cranial structures, the direct method and a photographic method, have been compared in day 18 mouse fetuses that have been stained with alizarin red S and cleared. Variation was determined for repeated measurements with each technique of the same specimen as well as the variation among specimens. There was variation in the results of direct measurements of the same specimens, but this was significantly (P < 0.5) less than the variation among specimens. By comparison, the variation of measurements of the same specimen made using the photographic method was not significantly (P > 0.2) different from the variation among the specimens, and the variation of the specimens was not significantly (P > 0.2) different between the photographic and direct methods. Direct measurement is recommended for making linear measurements on the cleared skull. The photograph method allows better delineation of angles and calculation of surface area.