Novel Multifunctional Hybrids of Single-Walled Carbon Nanotubes with Nucleic Acids: Synthesis and Interactions with Living Cells

Novel hybrids of fluorescein-labeled poly­(ethylene glycol)-modified single-walled carbon nanotubes (SWCNTs) with nucleic acids were prepared. 5′-Pyrene conjugates of oligodeoxyribonucleotides were used to construct the noncovalent hybrids, with the pyrene residues acting as anchor groups, immobilizing an oligonucleotide on the SWCNT surface. The hybrid formation characteristics were studied using ζ-potential measurements and adsorption isotherm plots. Transmission electron microscopy (TEM) of the samples stained with contrast agents proved that the pyrene conjugates of oligonucleotides were adsorbed onto the surfaces of the functionalized SWCNTs. On the basis of the MTT assay, the functionalized SWCNTs and their hybrids with oligonucleotides exhibited low toxicity toward HeLa, KB-3-1, and KB-8-5 cells. A TEM study of ultrathin sections of cells treated with SWCNTs revealed that the nanotubes directly interacted with the cellular surface

Appearance and size of urine EVs.

<p>(A) TEM images of EVs from urine of HD and PCa patients before and after 0.1 μm filtration. (B) Size distribution of EVs before and after 0.1 μm filtration. Mean size with error bars for range. Additional TEM images can be found in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0157566#pone.0157566.s001" target="_blank">S1 Fig</a>.</p

Total RNA in preparations of urine EVs.

<p>Bioanalyzer traces of total RNA from ERV fraction of the urine of HD (A) and PCa patient (B). The data from Agilent 2100 Bioanalyzer with 25 nt RNA fragment as an internal standard.</p

Example miR-19b correlation plots.

<p>Correlation plots of miR-19b concentration in TEV and ERV fractions of healthy individuals (A) and prostate cancer patients (B). AU—arbitrary units of concentration.</p

Overview of the study population.

<p>Overview of the study population.</p

Expression and diagnostic value of miRNA in HD and PCa patients.

<p>Analytical performance ROC analysis and statistical analysis of the results (non-parametric Mann-Whitney test) of miRNA expression in urine.</p

Protein concentration in urine EVs.

<p>Tukey box plots of concentration of total protein in preparations of EVs isolated from the urine of PCa patients and HD were determined using NanoOrange fluorescent dye and presented as mg per ml of probe (A) or ng per ml of urine (B).</p

Relative miRNA expression in urine EVs.

<p>Tukey box plots of miRNA expression in all fractions of urine. Asterisks (*) indicate statistical significance. *P<0.0001; **P = 0.0150.</p

Spearman’s correlation of miRNA expression between the fractions.

<p>Concentration was normalized to the starting urine volume.</p