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Nanostraw-Assisted Cellular Injection of Fluorescent Nanodiamonds via Direct Membrane Opening.

Small (Weinheim an der Bergstrasse, Germany) (2021-01-28)
Elke Hebisch, Martin Hjort, Diogo Volpati, Christelle N Prinz
ABSTRACT

Due to their stable fluorescence, biocompatibility, and amenability to functionalization, fluorescent nanodiamonds (FND) are promising materials for long term cell labeling and tracking. However, transporting them to the cytosol remains a major challenge, due to low internalization efficiencies and endosomal entrapment. Here, nanostraws in combination with low voltage electroporation pulses are used to achieve direct delivery of FND to the cytosol. The nanostraw delivery leads to efficient and rapid FND transport into cells compared to when incubating cells in a FND-containing medium. Moreover, whereas all internalized FND delivered by incubation end up in lysosomes, a significantly larger proportion of nanostraw-injected FND are in the cytosol, which opens up for using FND as cellular probes. Furthermore, in order to answer the long-standing question in the field of nano-biology regarding the state of the cell membrane on hollow nanostructures, live cell stimulated emission depletion (STED) microscopy is performed to image directly the state of the membrane on nanostraws. The time-lapse STED images reveal that the cell membrane opens entirely on top of nanostraws upon application of gentle electrical pulses, which supports the hypothesis that many FND are delivered directly to the cytosol, avoiding endocytosis and lysosomal entrapment.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Triton X-100, laboratory grade
Sigma-Aldrich
Penicillin-Streptomycin, Solution stabilized, with 10,000 units penicillin and 10 mg streptomycin/mL, 0.1 μm filtered, BioReagent, suitable for cell culture
Sigma-Aldrich
Paraformaldehyde, powder, 95%
Sigma-Aldrich
Bovine Serum Albumin, chromatographically purified, New Zealand origin, low endotoxin, suitable for cell culture, pH 7, ≥98%