Membrane Opening on Nanostraws Revealed By Live-Cell STED Microscopy Imaging

Nanotubes and nanostraws are increasingly used to transfect cells with better efficiencies and cell viabilities compared to existing transfection methods (liposomes, viral vectors and electroporation)^[1,2]. A long-standing question in the field concerns the state of the cell membrane on such hollow nanostructures. Here, in order to investigate the state of the cell membrane during and after transfection, we have performed video-rate live cell STED microscopy imaging of the cell membrane when interfaced with nanostraws. The time-lapse STED images reveal that the cell membrane opens entirely on top of nanostraws upon application of gentle electrical pulses, and that it recovers and seals 30-60 min after turning off the electrical field^[3]. We further demonstrate that we can achieve direct delivery of fluorescent nanodiamonds to the cytosol using nanostraws. The nanostraw delivery leads to efficient and rapid nanodiamond transport into cells compared to when incubating cells in nanodiamond-containing medium. Moreover, whereas all internalized nanodiamonds delivered by incubation end-up in lysosomes, a significant larger proportion of nanostraw-injected nanodiamonds are located in the cytosol, which opens up for using nanodiamonds as cellular probes<br/>References:<br/>[1] X. Xie, A. M. Xu, S. Leal-Ortiz, Y. Cao, C. C. Garner, N. A. Melosh, <i>ACS Nano</i> <b>2013</b>, <i>7</i>, 4351.<br/>[2] L. Schmiderer, A. Subramaniam, K. Zemaitis, A. Bäckström, D. Yudovich, S. Soboleva, R. Galeev, C. N. Prinz, J. Larsson, M. Hjort, <i>Proc. Natl. Acad. Sci. U. S. A.</i> <b>2020</b>, <i>117</i>, 21267.<br/>[3] E. Hebisch, M. Hjort, D. Volpati, C. N. Prinz, <i>Small</i> <b>2021</b>, <i>17</i>, 2006421.