Alex594 conjugated goat anti mouse or rabbit igg

For immunofluorescent staining, isolated cerebral tissues were mounted in OCT (optimal cutting temperature compound) and used for preparing cryosections. Cryosections (10 μm) were fixed and permeated with acetone or 4% paraformaldehyde with 0.5% Triton X-100, and then incubated with antibodies against Pnn, SRSF1, SRSF2, and p53bp1 at 4 °C overnight. Sections were then stained with Alex488- or Alex594-conjugated goat antimouse or rabbit IgG (Invitrogen). After counterstaining with DAPI, sections were examined under a fluorescent microscope (BX53, Olympus, Tokyo, Japan).

For immunofluorescent staining, isolated quadriceps were mounted in OCT and used for preparing cryosections. Sections were fixed and permeated with ice-cold acetone or 4% paraformaldehyde with 0.5% Triton X-100, and then incubated with antibodies against CD31 at 4 °C overnight. Coverslips or slides were then incubated with Alex594-conjugated goat anti-mouse or rabbit IgG (Invitrogen, Carlsbad, CA, USA). After counterstaining with DAPI, sections were examined under a fluorescent microscope.

For immunofluorescent staining, isolated myocardium was mounted in OCT and used for preparing cryosections, while cardiomyocytes were seeded on coverslips for 24 h. Sections or coverslips were fixed and permeated with ice-cold acetone or 4% paraformaldehyde with 0.5% Triton X-100, and then incubated with antibodies against CD90, Troponin I, CD68, or CD11b at 4 °C overnight. Coverslips or slides were then incubated with Alex488 or Alex594-conjugated goat anti-mouse or rabbit IgG (Invitrogen, Carlsbad, CA, USA). After counterstaining with DAPI, samples were examined under a fluorescent microscope. To analyze the extent of collagen synthesis and deposition, cardiac paraffin sections (6 µm) at 3 mm intervals were stained with Masson’s Trichrome. Masson’s Trichrome staining was performed according to the protocol provided by the manufacturer. In brief, sections were firstly fixed in Bouin’s solution. After incubation in Weigert’s Iron Hematoxylin solution, the slides were stained with Biebrich Scarlet-Acid Fuchsin and Aniline Blue, and then dehydrated in ethanol and xylene.