A Technique to Increase Accessibility to Late‐Stage Chick Embryos for In Ovo Manipulations

Abstract

Background During early development avian embryos are easily accessible in ovo for transplantations and experimental perturbations. However, these qualities of the avian embryonic model rapidly wane shortly after embryonic day (E)4 when the embryo is obscured by extraembryonic membranes, making it difficult to study developmental events that occur at later stages in vivo. Results and Conclusions In this study, we describe a multistep method that involves initially windowing eggs at E3, followed by dissecting away extraembryonic membranes at E5 to facilitate embryo accessibility in ovo until later stages of development. The majority of the embryos subjected to this technique remain exposed between E5 and E8, then become gradually displaced by the growing allantois from posterior to anterior regions. Exposed embryos are viable and compatible with embryological and modern developmental biology techniques including tissue grafting and ablation, gene manipulation by electroporation, and protein expression. This technique opens up new avenues for studying complex cellular interactions during organogenesis and can be further extrapolated to regeneration and stem cell studies.

Optimized <em>Ex-ovo</em> Culturing of Chick Embryos to Advanced Stages of Development

“…The above challenges in the ex ovo method has meant that researchers turn to the windowing method 9,10 to view embryonic chick development.…”

Utilizing Chick Embryo in Ovo Model System to Study the Effects of Misoprostol on Early Embryogenesis - A Pilot Study

“…Yalcin et al [19] have utilized ex ovo chick embryo culture system to study the relationships between genetic and microenvironmental factors driving normal and malformed embryonic development [19]. During the initial days of incubation, a chick embryo, which begins as a blastoderm undergoes gastrulation, neurulation and several morphogenetic movements to form distinct cranial and trunk structures [20]. These developmental features resemble closely to human development process.…”

A comparative analysis of chick culturing methods on skeletogenesis

“…de Gennaro (1980) hypothesized that a flat‐bottomed culturing dish, that restricts the chorioallantoic membrane from completely surrounding the embryo and impeding the embryo's ability to move freely, could cause abnormalities in the morphology of the endochondral elements. However, more recently, Spurlin and Lwigale () demonstrated that preventing the chorioallantoic membrane from completely surrounding the embryo in‐ovo does not grossly affect the morphology of the embryo. Therefore, we do not believe that the changes in chorioallantoic membrane position, due to shell‐less culturing in a flat‐bottomed dish, caused the abnormalities observed in the endochondral elements.…”

“…For each of these culturing methods, there are a few different techniques. In‐ovo culturing inside a windowed egg [can be conducted with the removal of albumin (Korn and Cramer, ; Spurlin and Lwigale, ) or without this removal (Speksnijder and Ivarie, )]. Additionally, this window culturing technique can utilize wax or an adhesive tape to seal the windows in the eggshells.…”