Universal intracellular biomolecule delivery with precise dosage control.
Universal intracellular biomolecule delivery with precise dosage control.
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DOI：
10.1126/sciadv.aat8131
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发表时间：
2018-10
期刊：
影响因子：
13.6
通讯作者：
Melosh NA
中科院分区：
文献类型：
--
作者：
Cao Y;Chen H;Qiu R;Hanna M;Ma E;Hjort M;Zhang A;Lewis RS;Wu JC;Melosh NA
我们展示了使用纳米管电穿孔将DNA、mRNA和蛋白质定量地控制递送到原代细胞中。mRNA、DNA和其他大分子的细胞内递送在一系列生物学研究和临床治疗中起着至关重要的作用。然而，目前的方法在递送的材料量方面产生了很大的变化，并且对细胞类型和可能的货物有限制。在这里，我们展示了定量控制交付到一系列的原代细胞和细胞系与紧密的剂量分布使用纳米管电穿孔系统（内斯）。在内斯中，细胞被培养在具有突出纳米管的径迹蚀刻膜上，所述突出纳米管连接到膜下方的流体环境。紧密的细胞-纳米管界面将施加的电场集中到细胞膜上，从而实现细胞膜的低电压和非损伤性局部穿孔。与此同时，磁场将生物分子通过纳米管注入细胞的同一位置。我们表明，所提供的材料的量是精确控制的所施加的电压，交付时间，和试剂浓度。内斯甚至对于原代细胞类型或不同的细胞密度也是高度有效的，在很大程度上是货物不可知的，并且可以同时递送特定比例的不同分子。使用简单的细胞培养孔格式，内斯在20秒内递送到> 100，000个细胞中，细胞活力>95%，使得能够容易地、剂量受控地进行细胞内递送，用于多种生物应用。
We demonstrate quantitatively controlled delivery of DNA, mRNA, and protein into primary cells using nanostraw electroporation. Intracellular delivery of mRNA, DNA, and other large macromolecules into cells plays an essential role in an array of biological research and clinical therapies. However, current methods yield a wide variation in the amount of material delivered, as well as limitations on the cell types and cargoes possible. Here, we demonstrate quantitatively controlled delivery into a range of primary cells and cell lines with a tight dosage distribution using a nanostraw-electroporation system (NES). In NES, cells are cultured onto track-etched membranes with protruding nanostraws that connect to the fluidic environment beneath the membrane. The tight cell-nanostraw interface focuses applied electric fields to the cell membrane, enabling low-voltage and nondamaging local poration of the cell membrane. Concurrently, the field electrophoretically injects biomolecular cargoes through the nanostraws and into the cell at the same location. We show that the amount of material delivered is precisely controlled by the applied voltage, delivery duration, and reagent concentration. NES is highly effective even for primary cell types or different cell densities, is largely cargo agnostic, and can simultaneously deliver specific ratios of different molecules. Using a simple cell culture well format, the NES delivers into >100,000 cells within 20 s with >95% cell viability, enabling facile, dosage-controlled intracellular delivery for a wide variety of biological applications.
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影响因子：
64.5
作者：
Park CY;Hoover PJ;Mullins FM;Bachhawat P;Covington ED;Raunser S;Walz T;Garcia KC;Dolmetsch RE;Lewis RS
通讯作者：
Lewis RS
影响因子：
48
作者：
Mali, Prashant;Esvelt, Kevin M.;Church, George M.
通讯作者：
Church, George M.
影响因子：
10.8
作者：
Kang W;Yavari F;Minary-Jolandan M;Giraldo-Vela JP;Safi A;McNaughton RL;Parpoil V;Espinosa HD
通讯作者：
Espinosa HD
影响因子：
4.6
作者：
Lönn P;Kacsinta AD;Cui XS;Hamil AS;Kaulich M;Gogoi K;Dowdy SF
通讯作者：
Dowdy SF
影响因子：
46.9
作者：
Hendel A;Bak RO;Clark JT;Kennedy AB;Ryan DE;Roy S;Steinfeld I;Lunstad BD;Kaiser RJ;Wilkens AB;Bacchetta R;Tsalenko A;Dellinger D;Bruhn L;Porteus MH
通讯作者：
Porteus MH