Immunofluorescent Imaging of Cardiac Tissue

Immunofluorescence was performed on formalin-fixed paraffin-embedded heart sections (5 µm). Briefly, the sections were deparaffinized, followed by antigen retrieval and blocked with blocking buffer (1% BSA in 1X PBS) for 1 h. Similarly, OCT-embedded sections were fixed with 4% paraformaldehyde for 20 min and rehydrated in PBS for 30 min. Then, sections were incubated overnight in a humidified chamber at 4°C with primary antibody against rat anti-mouse neutrophil (Serotec), rat anti-mouse F4/80 (Serotec), mouse anti-nitrotyrosine (Santa Cruz), mouse anti-4-HNE (Abcam). Finally, sections were incubated with different fluorophore-conjugated secondary antibodies (Invitrogen USA), as described previously (26 (link), 32 (link)).