A comparative analysis of chick culturing methods on skeletogenesis

Chick embryos are desirable models for the study of developmental biology. Despite this, there are very few studies that examine the effect of different culturing methods on skeletogenesis, specifically, intramembranous and endochondral bones. This study presents a detailed description of these effects by comparing two different culturing methods: windowed (in the shell) eggs and ex-ovo or shell-less culturing to normal development. Using whole mount bone staining, we determined that there is no significant difference in the length of the ossified region of intramembranous and endochondral bones in control versus window cultured embryos. However, these bones are significantly underossified in shell-less embryos. Shell-less embryos also exhibit abnormalities in endochondral bones. Intramembranous bones, interestingly, are morphologically normal in shell-less embryos. This study provides the first detailed description of ossification in window (in-ovo) and shell-less (ex-ovo) cultured embryos compared with controls (in-ovo). Patterning of the skeleton is unaffected regardless of culturing method. We conclude that studies involving endochondral bones should not utilise shell-less culturing methods. This data has been lacking in the literature and will serve as an important resource for those using cultured chick embryos in the study of skeletogenesis.