Interfacing silicon nanowires with mammalian cells.
نویسندگان
چکیده
Nanotechnology has received increased attention in the biological research field. The important examples are (1) the usage of nanoparticles in optical and magnetic resonance imaging;1,2 (2) the demonstration of potential application of metal nanoshells and carbon nanotubes for the treatment of tumor and cancer cells;3,4 and (3) the application of nanowire-based transistors to electrically detect specific biomolecules.5,6 In all of these cases, the nanomaterials are functioning either inside the cells or at the vicinity of the surface of biomolecules. Direct interconnection of the cells to the external world by interfacing nanomaterials may afford great opportunities to probe and manipulate biological processes occurring inside the cells, across the membranes, and between neighboring cells.7,8 A nanoscale material with high aspect ratio is a good candidate for this application. For instance, silicon nanowires (SiNWs, d ) 1-100 nm) are a few orders of magnitude smaller in diameter than mammalian cells (dcell ∼ on the order of 10 μm) yet comparable to the sizes of various intracellular biomolecules. The nanowires have high aspect ratio (<103) and yet are sufficiently rigid to be mechanically manipulated. The nanometer scale diameter and the high aspect ratio of SiNWs make them readily accessible to the interiors of living cells, which may facilitate the study of the complex regulatory and signaling patterns at the molecular level. In this Communication, we present the first demonstration of a direct interface of silicon nanowires with mammalian cells such as mouse embryonic stem (mES) cells and human embryonic kidney (HEK 293T) cells without any external force. The cells were cultured on a silicon (Si) substrate with a vertically aligned SiNW array on it. The penetration of the SiNW array into individual cells naturally occurred during the cell incubation. The cells survived up to several days on the nanowire substrates. The longevity of the cells was highly dependent on the diameter of SiNWs. Furthermore, successful maintenance of cardiac myocytes derived from mES cells on the wire array substrates was observed, and gene delivery using the SiNW array was demonstrated. SiNWs were synthesized vertically aligned with respect to Si(111) substrates via chemical vapor deposition as described earlier.9 The diameter of the nanowires was controlled by the size of gold nanoparticles that were used as catalytic seeds for the nanowire synthesis or by reducing the diameter of Si nanowires via oxidation and subsequent hydrofluoric (HF) acid etching step.10 The SiNW substrates had a native oxide layer and were used without any surface modification unless otherwise specified. Before any exposure to living cells, the substrates were sterilized in a solvent of 70% ethanol and 30% sterile water. First, physical interaction between the nanowires and the cells was studied using confocal microscopy and scanning electron microscopy (SEM). Mouse embryonic stem cells stably expressing green fluorescent protein (GFP) were cultured on silicon nanowire array substrates in standard non-differentiation medium. To facilitate visualization, a nanowire array was prepared with a high density (20-30 nanowires/cell). The diameter and the length of the nanowires were d ∼ 90 nm and L ∼ 6 μm, respectively. 50 000100 000 cells were incubated with a substrate deposited in a well of a 24-well plate. As the cells in the culture medium subsided on the nanowire substrate, the cells were penetrated by silicon nanowires as revealed by microscopy. Figure 1a shows a confocal microscopy image of the pierced cells. The nanowires appeared as black dots inside the cells. The focal plane was adjusted to the top of the nanowires inside the cells. Each dot corresponds to a local area of a cell that was occupied by each nanowire. SEM images of the samples are shown in Figure 1b-d. To maintain the morphology of the cells for SEM, the samples were prepared via a critical point drying technique after the treatment with glutaraldehyde for fixation and osmium tetroxide for contrast enhancement. Figure 1c,d shows individual cells penetrated by silicon nanowires. Several nanowires inside and underneath the cells can be clearly seen. The penetration was routinely observed within an hour of the incubation by fluorescent microscopy. No external force was necessary for the penetration, owing to the small diameter and high aspect ratio of nanowires. The results indicate that nanowires can be readily introduced inside the cells. Next, viability of the penetrated cells was studied via the proliferation of the cells and propidium iodide (PI) staining. To ensure that we study only the cells that were pierced with nanowires, the substrates were first placed in a culture medium containing cells for an hour for the cell subsidence and then transferred into fresh medium without cells for further incubation. The density of † Lawrence Berkeley National Laboratory, University of California, Berkeley. ‡ Department of Chemistry, University of California, Berkeley. § Gladstone Institute of Cardiovascular Disease. Figure 1. (a) A confocal microscopy image of mouse embryonic stem (mES) cells penetrated with silicon nanowires. (b) SEM image of mES cells on a nanowire array substrate. (c, d) SEM images of individual mES cells penetrated with silicon nanowires. The diameter and the length of the nanowires are ∼90 nm and ∼6 μm, respectively. Published on Web 05/22/2007
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ورودعنوان ژورنال:
- Journal of the American Chemical Society
دوره 129 23 شماره
صفحات -
تاریخ انتشار 2007