2.12 Histological analysis of skin tissue
The RS-NC-injected mouse skin
tissue was observed by histological analysis after H&E staining. All
animal experiments were performed according to the protocol approved by
the Institutional Animal Care and Use Committee (IACUC) of Incheon
National University (INU-ANIM-2021-01). First, RS-NC (N/P ratio 5)
containing 200 pmol of negative control siRNA were incubated for 30
minutes at RT. 30 µL of RS-NC and DPBS as negative control were injected
intradermally to epilated back of each five-week-old BALB/c mouse
(Orientbio, Inc., Seongnam, South Korea). After 4 days, the injected
skin tissues of the euthanized mice were excised and fixed in 10% (w/v)
formalin (Sigma-Aldrich, MO, USA). After 22 hours, the tissue samples
were dehydrated, paraffin-embedded by embedding center (Histostar;
Thermo Fisher Scientific, MA, USA), and finally sectioned to 4 µm
thickness by microtome (HM 355S; Thermo Fisher Scientific, MA, USA), and
placed on the slide. After deparaffinization, the tissue samples were
stained with hematoxylin and eosin (BioGnost, Zagreb, Croatia). Then,
the slides were observed with Leica DM1000 microscope (Leica
Microsystems, Hesse, Germany) under 20× magnification.