Department of Biomedical Engineering, Thoraxcenter, Erasmus MC University Medical Center Rotterdam;
Department of Biomedical Engineering, Thoraxcenter, Erasmus MC University Medical Center Rotterdam.
J Vis Exp. 2021 Feb 9(168). doi: 10.3791/62076.
The chicken embryo and the blood-vessel rich chorioallantoic membrane (CAM) is a valuable in vivo model to investigate biomedical processes, new ultrasound pulsing schemes, or novel transducers for contrast-enhanced ultrasound imaging and microbubble-mediated drug delivery. The reasons for this are the accessibility of the embryo and vessel network of the CAM as well as the low costs of the model. An important step to get access to the embryo and CAM vessels is to take the egg content out of the eggshell. In this protocol, three methods for taking the content out of the eggshell between day 5 and 8 of incubation are described thus allowing the embryos to develop inside the eggshell up to these days. The described methods only require simple tools and equipment and yield a higher survival success rate of 90% for 5-day, 75% for 6-day, 50% for 7-day, and 60% for 8-day old incubated eggs in comparison to ex ovo cultured embryos (~50%). The protocol also describes how to inject cavitation nuclei, such as microbubbles, into the CAM vascular system, how to separate the membrane containing the embryo and CAM from the rest of the egg content for optically transparent studies, and how to use the chicken embryo and CAM in a variety of short-term ultrasound experiments. The in vivo chicken embryo and CAM model is extremely relevant to investigate novel imaging protocols, ultrasound contrast agents, and ultrasound pulsing schemes for contrast-enhanced ultrasound imaging, and to unravel the mechanisms of ultrasound-mediated drug delivery.
鸡胚和富含血管的绒毛尿囊膜（CAM）是一种非常有价值的体内模型，可用于研究生物医学过程、新的超声脉冲方案，或用于对比增强超声成像和微泡介导药物输送的新型换能器。其原因在于胚胎和 CAM 血管网络的可及性以及该模型的低成本。进入胚胎和 CAM 血管的一个重要步骤是将卵内容物从蛋壳中取出。在本方案中，描述了在孵育的第 5 至第 8 天之间从蛋壳中取出卵内容物的三种方法，从而使胚胎在蛋壳内发育至这些天。所描述的方法仅需要简单的工具和设备，与体外培养的胚胎（约 50%）相比，5 天、6 天、7 天和 8 天龄孵育鸡蛋的胚胎成活率更高，分别为 90%、75%、50%和 60%。该方案还描述了如何将空化核（如微泡）注入 CAM 血管系统，如何将含有胚胎和 CAM 的膜与卵内容物的其余部分分离以进行光学透明研究，以及如何在各种短期超声实验中使用鸡胚和 CAM。体内鸡胚和 CAM 模型非常适合研究新型成像方案、超声对比剂和超声脉冲方案，以用于对比增强超声成像，并揭示超声介导药物输送的机制。
