Gold Nanorod Induced Warming of Embryos from the Cryogenic State Enhances Viability
Gold Nanorod Induced Warming of Embryos from the Cryogenic State Enhances Viability
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DOI：
10.1021/acsnano.7b02216
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发表时间：
2017-08-01
期刊：
影响因子：
17.1
通讯作者：
Bischof, John
中科院分区：
文献类型：
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作者：
Khosla, Kanav;Wang, Yiru;Bischof, John
通过微量注射冷冻保护剂并快速冷却，斑马鱼胚胎可以达到稳定的低温状态，但胚胎的巨大尺寸一直导致在对流加热过程中失败。在这里，我们通过使用金纳米棒（gnr）来帮助升温过程来解决斑马鱼的低温保存问题。具体来说，我们将冷冻保护剂丙二醇与gnr一起微注射到斑马鱼胚胎中，并将样品在液氮中以90000℃/min的速率冷却。我们证明了在gnr激发下，用1064 nm激光脉冲照射1 ms的样品，可以快速解冻斑马鱼（1.4 X 10(7)℃/min）。这种快速的变暖过程导致了冰的形成，这可能会损害胚胎。14个试验（n = 223）的结果表明，可存活的胚胎在加热后1小时结构一致（31%），3小时继续发育（17%），24小时运动（10%）。这与对流加热对照组在所有时间点的活力、结构或运动为0%形成鲜明对比（n = 50, p < 0.001，方差分析）。我们基于纳米颗粒的加热过程可以应用于鱼类的储存，并且经过适当的修改，可以潜在地用于其他脊椎动物胚胎。
Zebrafish embryos can attain a stable cryogenic state by microinjection of cryoprotectants followed by rapid cooling, but the massive size of the embryo has consistently led to failure during the convective warming process. Here we address this zebrafish cryopreservation problem by using gold nanorods (GNRs) to assist in the warming process. Specifically, we microinjected the cryoprotectant propylene glycol into zebrafish embryos along with GNRs, and the samples were cooled at a rate of 90 000 degrees C/min in liquid nitrogen. We demonstrated the ability to unfreeze the zebrafish rapidly (1.4 X 10(7) degrees C/min) by irradiating the sample with a 1064 nm laser pulse for 1 ms due to the excitation of GNRs. This rapid warming process led to the outrunning of ice formation, which can damage the embryos. The results from 14 trials (n = 223) demonstrated viable embryos with consistent structure at 1 h (31%) and continuing development at 3 h (17%) and movement at 24 h (10%) postwarming. This compares starkly with 0% viability, structure, or movement at all time points in convectively warmed controls (n = 50, p < 0.001, ANOVA). Our nanoparticle-based warming process could be applied to the storage of fish, and with proper modification, can potentially be used for other vertebrate embryos.