A method to calibrate fetal μCT scans using histological sections from the same specimens

Craniofacial morphogenesis is an intricate developmental process in 3D, which therefore merits visualization and investigation in 3D. To better understand the process, we utilize μCT imaging, and describe a method to calibrate each cone beam μCT individually. Calibration is necessary, because during development, fetuses undergo tissue differentiation, which affects the acquisition process for radiographic images. Additionally, tissue fixation and conservation agents may influence the physical properties of the specimens and may affect image acquisition. After taking a μCT scan from each specimen, we separated a horizontal slice from each neck (which is inconsequential to our question with relation to the whole head). These neck specimens were prepared as horizontal histological serial sections and stained. With these as a reference, the μCT visualization parameters could be adjusted until they matched the selected virtual section planes, which correspond exactly to the planes of the histological sections with a precision (pixel size) of 0.69μm.