Survival rates of chick embryos cultured under different conditions.

<p>(a) Effect of the environment change. The survivability of chick embryos in the normal eggshell was 100%. The survivability of chick embryos in cubic eggshells fabricated using 0.1-mm-thick PDMS membranes (90%) was higher than that of the conventional culture method (71%). The chick embryos cultured in cubic eggshells without oxygen permeation died soon after transfer. (b) Effect of the thickness change. On day 7, the survivability of chick embryos in cubic eggshells fabricated using 0.3- and 0.5-mm-thick PDMS membranes was approximately 80% and 22%, respectively. Chick embryos in eggshells using 0.7-, 1.0-, and 1.3-mm-thick membranes died soon after transfer. This tendency is consistent with the oxygen permeation rate calculated using Equation (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118624#pone.0118624.e001" target="_blank">1</a>). (c) Heart rates of the chick embryos in normal eggshells and cubic eggshells fabricated using 0.1-, 0.3-, and 0.7-mm-thick membranes. Heart rates of chick embryos in cubic eggshells fabricated using 0.1-mm-thick membranes increased from 198 ± 6 to 239 ± 16 beats per minute, and eggshells using 0.3-mm-thick membranes increased from 188 ± 5 to 250 ± 9 beats per minute. The results were similar to the heart beats of chick embryos in normal eggshells. On the other hand, the average heart rates of chick embryos in eggshells using 0.7-mm-thick membranes decreased dramatically from day 5 because of the death of some chick embryos, which resulted in a high variance. The error bar is SEM (n = 5).