Departament de Biologia Cellular, Fisiologia i Immunologia, Facultat de Biociències, Universitat Autònoma de Barcelona, Bellaterra, Spain.
Hum Reprod. 2011 Jan;26(1):96-105. doi: 10.1093/humrep/deq309. Epub 2010 Nov 18.
Measures to prevent assisted reproductive technologies (ART) mix-ups, such as labeling of all labware and double-witnessing protocols, are currently in place in fertility clinics worldwide. Technological solutions for electronic witnessing are also being developed. However, none of these solutions eliminate the risk of identification errors, because gametes and embryos must be transferred between containers several times during an ART cycle. Thus, the objective of this study was to provide a proof of concept for a direct embryo labeling system using silicon-based barcodes.
Three different types of silicon-based barcodes (A, B and C) were designed and manufactured, and microinjected into the perivitelline space of mouse pronuclear embryos (one to four barcodes per embryo). Embryos were cultured in vitro until the blastocyst stage, and rates of embryo development, retention of the barcodes in the perivitelline space and embryo identification were assessed every 24 h. Release of the barcodes after embryo hatching was also determined. Finally, embryos microinjected with barcodes were frozen and thawed at the 2-cell stage to test the validity of the system after cryopreservation.
Barcodes present in the perivitelline space, independently of their type and number, did not affect embryo development rates. The majority of embryos (>90%) retained at least one of the microinjected barcodes in their perivitelline space up to the blastocyst stage. Increasing the number of barcodes per embryo resulted in a significant increase in embryo identification rates, but a significant decrease in the barcode release rates after embryo hatching. The highest rates of successful embryo identification (97%) were achieved with the microinjection of four type C barcodes, and were not affected by cryopreservation.
Our results demonstrate the feasibility of a direct embryo labeling system and constitute the starting point in the development of such systems.
防止辅助生殖技术（ART）混淆的措施，如对所有实验室器皿进行标记和双见证协议，目前已在世界各地的生育诊所实施。也正在开发用于电子见证的技术解决方案。然而，这些解决方案都不能消除身份识别错误的风险，因为配子和胚胎在一个 ART 周期中必须在容器之间多次转移。因此，本研究的目的是提供使用硅基条形码的直接胚胎标记系统的概念验证。
设计和制造了三种不同类型的硅基条形码（A、B 和 C），并将其微注射到小鼠原核胚胎的卵周隙中（每个胚胎一到四个条形码）。胚胎在体外培养至囊胚阶段，并在每 24 小时评估胚胎发育率、条形码在卵周隙中的保留率和胚胎识别率。还确定了条形码在胚胎孵化后的释放情况。最后，对注射有条形码的胚胎进行冷冻和解冻，在 2 细胞阶段测试系统在冷冻保存后的有效性。
无论条形码的类型和数量如何，存在于卵周隙中的条形码都不会影响胚胎发育率。大多数胚胎（>90%）在囊胚阶段之前至少保留了一个微注射的条形码。每个胚胎注射的条形码数量增加会导致胚胎识别率显著提高，但条形码在胚胎孵化后的释放率显著降低。注射四个 C 型条形码可获得最高的胚胎识别成功率（97%），且不受冷冻保存的影响。
我们的结果证明了直接胚胎标记系统的可行性，为该系统的开发奠定了基础。
