Optical tweezer-assisted cell pairing and fusion for somatic cell nuclear transfer within an open microchannel†

IF 6.1 2区 工程技术 Q1 BIOCHEMICAL RESEARCH METHODS Lab on a Chip Pub Date : 2024-11-06 DOI:10.1039/D4LC00561A
Yidi Zhang, Han Zhao, Zhenlin Chen, Zhen Liu, Hanjin Huang, Yun Qu, Yaowei Liu, Mingzhu Sun, Dong Sun and Xin Zhao
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Abstract

Somatic cell nuclear transfer (SCNT), referred to as somatic cell cloning, is a pivotal biotechnological technique utilized across various applications. Although robotic SCNT is currently available, the subsequent oocyte electrical activation/reconstructed embryo electrofusion is still manually completed by skilled operators, presenting challenges in efficient manipulation due to the uncontrollable positioning of the reconstructed embryo. This study introduces a robotic SCNT-electrofusion system to enable high-precision batch SCNT cloning. The proposed system integrates optical tweezers and microfluidic technologies. An optical tweezer is employed to facilitate somatic cells in precisely reaching the fusion site, and a specific polydimethylsiloxane (PDMS) chip is designed to assist in positioning and pairing oocytes and somatic cells. Enhancement in the electric field distribution between two parallel electrodes by PDMS pillars significantly reduces the required external voltage for electrofusion/electrical activation. We employed porcine oocytes and porcine fetal fibroblasts for SCNT experiments. The experimental results show that 90.56% of oocytes successfully paired with somatic cells to form reconstructed embryos, 76.43% of the reconstructed embryos successfully fused, and 70.55% of these embryos underwent cleavage. It demonstrates that the present system achieves the robotic implementation of oocyte electrical activation/reconstructed embryo electrofusion. By leveraging the advantages of batch operations using microfluidics, it proposes an innovative robotic cloning procedure that scales embryo cloning.

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开放式微通道内体细胞核移植的光镊辅助细胞配对与融合。
体细胞核移植(SCNT),又称体细胞克隆,是一项重要的生物技术,应用范围广泛。虽然目前已有机器人 SCNT,但随后的卵母细胞电激活/重建胚胎电融合仍需熟练操作人员手动完成,由于重建胚胎的定位不可控,给高效操作带来了挑战。本研究介绍了一种机器人 SCNT 电融合系统,以实现高精度批量 SCNT 克隆。该系统集成了光学镊子和微流控技术。光学镊子可帮助体细胞精确到达融合点,而特制的聚二甲基硅氧烷(PDMS)芯片可帮助卵母细胞和体细胞定位和配对。PDMS 柱增强了两个平行电极之间的电场分布,大大降低了电融合/电激活所需的外部电压。我们采用猪卵母细胞和猪胎儿成纤维细胞进行 SCNT 实验。实验结果表明,90.56% 的卵母细胞成功与体细胞配对形成重建胚胎,76.43% 的重建胚胎成功融合,70.55% 的胚胎发生裂殖。这表明,本系统实现了卵母细胞电激活/重建胚胎电融合的机器人操作。通过利用微流体技术批量操作的优势,该研究提出了一种创新的机器人克隆程序,从而扩大了胚胎克隆的规模。
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来源期刊
Lab on a Chip
Lab on a Chip 工程技术-化学综合
CiteScore
11.10
自引率
8.20%
发文量
434
审稿时长
2.6 months
期刊介绍: Lab on a Chip is the premiere journal that publishes cutting-edge research in the field of miniaturization. By their very nature, microfluidic/nanofluidic/miniaturized systems are at the intersection of disciplines, spanning fundamental research to high-end application, which is reflected by the broad readership of the journal. Lab on a Chip publishes two types of papers on original research: full-length research papers and communications. Papers should demonstrate innovations, which can come from technical advancements or applications addressing pressing needs in globally important areas. The journal also publishes Comments, Reviews, and Perspectives.
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