Open Access Open Badges Methodology article

Surface landmark quantification of embryonic mouse craniofacial morphogenesis

Christopher J Percival1, Rebecca Green2, Ralph Marcucio3 and Benedikt Hallgrímsson1*

Author Affiliations

1 Department of Cell Biology and Anatomy, Alberta Children’s Hospital Institute for Child and Maternal Health, The McCaig Bone and Joint Institute, University of Calgary, Calgary, AB T2N 4N1, Canada

2 Department of Craniofacial Biology and Program in Reproductive Sciences, University of Colorado – Denver, Denver, CO 80045, USA

3 Department of Orthopaedic Surgery, The Orthopaedic Trauma Institute, UCSF School of Medicine, San Francisco, CA 94143, USA

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BMC Developmental Biology 2014, 14:31  doi:10.1186/1471-213X-14-31

Published: 24 July 2014



Morphometric quantification of subtle craniofacial variation in studies of experimentally modified embryonic mice has proved valuable in determining the effects of developmental perturbations on craniofacial morphogenesis. The direct comparison of landmark coordinate data from embryos of many different mouse strains and mouse models can advance our understanding of the bases for craniofacial variation. We propose a standard set of craniofacial surface landmarks, for use with embryonic day (E) 10.5-12.5 mice, to serve as the foundation for this type of data compilation and analysis. We quantify the intra- and inter-observer landmark placement variation associated with each landmark and determine how the results of a simple ontogenetic analysis might be influenced by selection of landmark set.


Intraobserver landmark placement error for experienced landmarkers generally remains below 0.1 mm, with some landmarks exhibiting higher values at E11.5 and E12.5. Interobserver error tends to increase with embryonic age and those landmarks defined on wide inflections of curves or facial processes exhibit the highest error. Landmarks with highest intra- or inter-observer are identified and we determine that their removal from the dataset does not significantly change the vectors of craniofacial shape change associated with an ontogenetic regression.


Our quantification of landmark placement error demonstrates that it is preferable for a single observer to identify all landmark coordinates within a single study and that significant training and experience are necessary before a landmarker can produce data for use in larger meta-analyses. However, we are confident that this standard landmark set, once landmarks with higher error are removed, can serve as a foundation for a comparative dataset of facial morphogenesis across various mouse populations to help identify the developmental bases for phenotypic variation in the craniofacial complex.

Morphometrics; Landmark error; Facial prominences; Craniofacial morphogenesis; Mouse embryo; 3D imaging; Micro-CT