Alexa fluor 647 anti mouse cd106 vcam 1 antibody

Ten-micrometer sections of frozen heart tissue were incubated at room temperature with primary anti–IL-36R, anti–IL-36α, anti–IL-36β, or IgG control antibodies (1:100 dilution; R&D Systems, polyclonal) and a secondary donkey anti–goat Alexa Fluor 488 antibody (1:100 dilution; Abcam, polyclonal). Sections were also incubated with a PE anti–mouse CD31 antibody (1:100 dilution, BioLegend, clone 390), an Alexa Fluor 647 anti–mouse CD106/VCAM-1 antibody (1:100 dilution, BioLegend, clone 429), or an anti-DNA/RNA damage antibody to detect oxidative damage (1:100 dilution, Abcam, clone 153A). Images were captured using an EVOS FL (Thermo Fisher Scientific) or multiphoton microscope (FVMPE-RS, Olympus). ImageJ was used to quantify the MFI of each image with additional analysis of MFI on regions containing only coronary capillaries or a large blood vessel.

Frozen heart tissue sections (10 µm) were incubated with primary antibodies against IL-36R, IL-36α, IL-36β, or IgG control antibodies (1:100 dilution; R & D Systems) and a secondary donkey anti-goat Alexa Fluor-488 antibody (1:100 dilution; Abcam) at room temperature. Sections were also incubated with PE anti-mouse CD31 antibody (1:100 dilution, Biolegend), Alexa Fluor-647 anti-mouse CD106/VCAM-1 antibody (1:100 dilution, Biolegend), or an anti-DNA/RNA damage antibody for oxidative damage detection (1:100 dilution, Abcam). Images were captured using an EVOS FL microscope (ThermoFisher Scientific), and ImageJ was used to quantify mean fluorescence intensity (MFI) for each image.