Micromachined Capillary Arrays for DNA Injection

本研究的目的是利用微机械加工技术制备用于活细胞DNA注射的微毛细管阵列。注射装置由规则排列的毛细血管组成。毛细血管被推到细胞上并刺穿细胞。像DNA这样的遗传物质可以通过中空的毛细血管注入细胞。毛细血管通常直径为1-5微米，厚度为0.5-1微米，高度为20-50微米，它们之间的间距为几十微米。主要研究成果有：(1)建立了毛细管阵列的微加工工艺。采用电感耦合等离子体反应离子刻蚀技术在硅衬底上刻蚀出直径为5微米、深度为100 ~ 200微米的小孔。然后对孔内表面进行热氧化处理（厚度0.5-1微米）。从背面蚀刻硅衬底露出底孔。通过缓冲HF的短刻蚀去除底部的氧化层，形成一个开放的毛细管。进一步蚀刻硅衬底使毛细血管突出20-30微米的高度。最后，在顶部表面阳极粘合一个玻璃盖，作为储液器和连接管道和泵的外部连接器。(2)将所制备的装置与培养的烟草细胞进行对照。DNA溶液通过穿刺毛细血管注射。我们确认细胞中产生了一种编码酶。(3)我们还制造了一种细胞捕获装置，使细胞保持在与注射装置相同间距的规则阵列中。该装置的通孔比电池直径更细。悬浮在液体中的细胞通过从背面吸液体而被捕获在孔上。

The purpose of this research is to fabricate micro capillary arrays for DNA injection to living cells by using micromachnining technology. The injection device is composed of the regular array of capillaries. Capillaries were pushed against cells and pierced them. Genetic substances such as DNA can be injected into cells through hollow capillaries. Capillaries are typically 1-5 micrometers in diameter, 0.5-1 micrometers in thickness, 20-50 micrometers in height and a few tens of micrometers in pitch between them. Major achievements of the research are :(1) We established the micromachining process for the capillary array. Thin holes of 5 micrometers in diameter and 100-200 micrometers in depth were etched in the silicon substrate by ICP-RIE (inductive coupled plasma-reactive ion etching). Then the inner surface of holes was thermally oxidized (0.5-1 micrometers in thickness). Etching of the silicon substrate from the backside revealed the bottom holes. The oxide layer at the bottom was removed by the short etching of buffered HF to make an open capillary. Further etching of the silicon substrate made capillaries protrude for 20-30 micrometers in height. Finally, a glass cover was anodically bonded on the top surface serving as a liquid reservoir and an external connector to a pipe and a pump.(2) The fabricated device was set against cultured tobacco cells. DNA solution was injected through piercing capillaries. We confirmed the production of a coded enzyme in cells.(3) We also fabricated a cell-capturing device to hold cells in a regular array with the same pitch of the injection device. The device had thinner through holes than the cell diameter. Cells suspended in liquid were captured on the holes by sucking the liquid through them from the backside.