*2.13. Confocal Fluorescence Microscopy*

For confocal microscopy, HepG2 cells cultivated in 1 cm<sup>2</sup> chambers slide (Lab-Tek, Thermo Fisher Scientific Inc., Waltham, MA, USA) were transfected with conjugate FAM-D-17PG or duplex D-17PG/FAM-17 to the final concentration of 5 μM in the media with various composition (see Section 2.11). Right after transfection, cells were washed twice with PBS to eliminate fluorescently labeled unbound oligonucleotides. Washed cell were fixed with 4% paraformaldehyde in DMEM for 30 min and washed twice with PBS. After removing the chamber, the microscopic slide containing cells was mounted with the cover slip glass using ProLong Gold antifade reagent with DAPI (Life Technologies, Eugene, OR, USA), then the slide was kept 24 h in the dark at room temperature. LSM 710 confocal microscope (Carl Zeiss Microscopy GmbH, Jena, Germany) was used in conjunction with Zen imaging software, and images were acquired with a Zeiss 63×/1.40 oil immersion objective. The excitation/emission laser wavelengths were 405 nm (to detect cell nuclei stained with DAPI), 488 nm (to detect FAM).
