Intracellular delivery and fate of peptide-capped gold nanoparticles

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TAT and HA2 Facilitate Cellular Uptake of Gold Nanoparticles but Do Not Lead to Cytosolic Localisation.

International audienceThe methods currently available to deliver functional labels and drugs to the cell cytosol are inefficient and this constitutes a major obstacle to cell biology (delivery of sensors and imaging probes) and therapy (drug access to the cell internal machinery). As cell membranes are impermeable to most molecular cargos, viral peptides have been used to bolster their internalisation through endocytosis and help their release to the cytosol by bursting the endosomal vesicles. However, conflicting results have been reported on the extent of the cytosolic delivery achieved. To evaluate their potential, we used gold nanoparticles as model cargos and systematically assessed how the functionalisation of their surface by either or both of the viral peptides TAT and HA2 influenced their intracellular delivery. We evaluated the number of gold nanoparticles present in cells after internalisation using photo-thermal microscopy and their subcellular localisation by electron microscopy. While their uptake increased when the TAT and/or HA2 viral peptides were present on their surface, we did not observe a significant cytosolic delivery of the gold nanoparticles

CCALNN-PEG monolayer proportion influence on gold nanoparticles uptake (Figure S3 dataset)

<p>HeLa cells were incubated in suspension with 5 nm diameter gold nanoparticles (final concentration 100 nM) coated with a mix of CALNN and CCALNN-PEG peptides for 10 min in serum-free medium and a further 30 min in complete medium (10 % FCS). Nanoparticles/medium were then discarded, cells transferred to a dish and left to attach for 4 h in complete medium, fixed and later imaged by photothermal microscopy.</p> <p>Images are two-plane TIFF images of the nanoparticles uptake by HeLa cells. The first image plane is the brightfield image, and the second image plane is the photothermal image.</p> <p>0%_CCALNN-PEG_&_100%_CALNN_[1-5].tif are images of cells incubated with nanoparticles coated with 100% CALNN peptides.<br>10%_CCALNN-PEG_&_90%_CALNN_[1-4].tif are images of cells incubated with nanoparticles coated with 90% CALNN and 10% CCALNN–PEG peptides.<br>20%_CCALNN-PEG_&_80%_CALNN_[1-4].tif are images of cells incubated with nanoparticles coated with 80% CALNN and 20% CCALNN–PEG peptides.<br>30%_CCALNN-PEG_&_70%_CALNN_[1-5].tif are images of cells incubated with nanoparticles coated with 70% CALNN and 30% CCALNN–PEG peptides.</p

Intracellular delivery of gold nanoparticles assisted by monolayer functionalisation with HA2/TAT peptides (Figure 1 dataset)

<p>HeLa cells were incubated with or without nanoparticles for 3h, fixed and imaged by photothermal microscopy.<br>When present nanoparticles were of a 5 nm diameter at a concentration of 500 nM.</p> <p>Images are two-plane TIFF images of the nanoparticles uptake by HeLa cells. The first image plane is the brightfield image, and the second image plane is the photothermal image.</p> <p>Cells_only_[1-4].tif are images of cells incubated without nanoparticles.<br>100%_capping_peptides_[1-4].tif are images of cells incubated with nanoparticles coated with a 4:1 ratio of CALNN : CCALNN-PEG peptides (capping peptides).<br>10%_dHA2_[1-4].tif are images of cells incubated with nanoparticles coated with 10 % CCALNN–dHA2 peptides and 90% capping peptides.<br>1%_Tat_[1-3].tif are images of cells incubated with nanoparticles coated with 1 % CALNN–TAT peptides and 99% capping peptides.<br>10%_dHA2_&_1%_Tat_[1-4].tif are images of cells incubated with nanoparticles coated with 10 % CCALNN–dHA2 peptides, 1 % CALNN–TAT peptides and 89% capping peptides.</p

Fmoc-diphenylalanine hydrogels: understanding the variability in reported mechanical properties

Fmoc-diphenylalanine (FmocFF or FmocPhePhe) is an important low molecular weight hydrogelator. Gelation can be induced by either lowering the pH of an aqueous solution of FmocFF or by the addition of water to a solution of FmocFF in a solvent such as DMSO. Despite the volume of literature on FmocFF, the mechanical properties reported for the gels vary significantly over four orders of magnitude and the origins of this variability is unclear. Here, we study systematically the mechanical properties of FmocFF gels prepared with different protocols. We demonstrate that the final pH of the gels is the principal determinant of the mechanical properties independently of the method of gel formation. We also show that additional variability arises from experimental factors such as the fraction of DMSO or the nature of the buffers used in selected systems

Nouvelle lecture des phases de la dernière déglaciation dans le Loch Sunart (Nord-Ouest Ecosse) basée sur des traceurs géochimiques

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Influence of the proportion of CCALNN-dHA2 peptides in the monolayer of gold nanoparticles.

<p>HeLa cells were incubated for 4h with 10nm gold nanoparticles (6nM) coated with a monolayer composed of two different proportions of the peptides CCALNN-dHA2 and CALNN, washed thoroughly with PBS, fixed and imaged by electron microscopy. (A-B) TEM images of the cells incubated with nanoparticle with the following peptide monolayer composition: (A) 50% CCALNN-dHA2 and 50% CALNN, (B) 100% CCALNN-dHA2. Arrowheads point toward gold nanoparticles either interacting with vesicular membranes (white) or displaying a cytosolic localisation (black). (C) Density of nanoparticles in endosomes estimated from images shown in (A) (n = 10, middle) and (B) (n = 10, right); quantification for 10% CCALNN-dHA2 (left) is the same as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0121683#pone.0121683.g002" target="_blank">Fig. 2E</a> (left). The sets of images analysed are available on figshare (<a href="http://dx.doi.org/10.6084/m9.figshare.875545" target="_blank">10.6084/m9.figshare.875545</a>, <a href="http://dx.doi.org/10.6084/m9.figshare.875477" target="_blank">10.6084/m9.figshare.875477</a>).</p

Nomenclature of the peptides utilised, with the short names and corresponding amino acid sequences.

<p>Nomenclature of the peptides utilised, with the short names and corresponding amino acid sequences.</p