抗冻DNA适体在人红细胞冷冻保存中的潜在应用

J. Bruno
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引用次数: 0

摘要

本文总结了概念验证实验,这些实验清楚地证明了针对早期冰晶核分子模拟物选择的DNA适体在缓慢冻融循环后保护人类红细胞完整性的能力。经过10个周期的筛选和DNA扩增,将罕见的候选DNA适体与铜-有机配体复合物(将水分子保持在类似早期冰晶核的构象中)相结合,通过相对比显微镜测试适体与不含适体的对照和与原始随机适体DNA文库模板一起,在-20°C慢速冷冻和过夜,在25°C慢速解冻后保存红细胞形态的能力。这些实验表明,最终选择的适体DNA分子库至少需要32 μg/ml才能完全保护几乎100%的红细胞。相比之下,没有适配体或随机适配体模板DNA(浓度为32 μg/ml)的处理组只产生了碎片化的红细胞和细胞碎片(没有完整的细胞),这表明需要特定选择的DNA构象来结合并限制形成冰晶的大小以冷冻保护红细胞。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Potential Use of Antifreeze DNA Aptamers for the Cryopreservation of Human Erythrocytes
This article summarizes proof of concept experiments which clearly demonstrated the ability of DNA aptamers selected against a molecular mimic of early ice crystal nuclei to protect the integrity of human erythrocytes following a slow freeze-thaw cycle. Following 10 cycles of selection and DNA amplification of rare candidate DNA aptamers against a copper-organic ligand complex which holds water molecules in a conformation resembling early ice crystal nuclei, the aptamers were tested for their ability to preserve erythrocyte morphology by phase-contrast microscopy versus controls without aptamers and with the original randomized aptamer DNA library template following slow freezing and storage overnight at -20 °C with slow thawing at 25 °C. Those experiments revealed that a minimum of 32 μg/ml of the final selected aptamer pool of DNA molecules was required to completely protect nearly 100% of the erythrocytes. By contrast, the treatment groups without the aptamers or with the randomized aptamer template DNA at 32 μg/ml produced only fragmented erythrocytes and cellular debris (no intact cells), thus indicating that specifically selected DNA conformations were required to bind and limit the size of forming ice crystals to cryoprotect the erythrocytes.
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