Mouse monoclonal alpha smooth muscle actin antibody

The lower body of postnatal day 0 (P0) or E11.5 mice were fixed overnight in 4% paraformaldehyde, paraffin-embedded, and sectioned at 6 μm for hematoxylin and eosin (H&E) staining or immunohistochemistry. For immunohistochemistry, tissues were deparaffinized and rehydrated. Antigen retrieval was performed by boiling the sections in 0.1 M sodium citrate buffer for 30 minutes. Samples were blocked for 30 minutes in PBST with 10% FBS. Sections were incubated with primary antibody overnight at 4°C. The primary antibodies used were mouse monoclonal anti-E-cadherin (BD Biosciences, 1:100, Franklin Lakes, NJ), mouse monoclonal calbindin-D-28K antibody (Sigma-Aldrich, 1:100), rabbit polyclonal cleaved caspase-3 antibody (Cell Signaling Technology, 1:100), and/or mouse monoclonal alpha smooth muscle actin antibody (Thermo Fisher Scientific, 1:100). Sections were washed with PBS and incubated at 4°C overnight with secondary antibodies, 488 nm conjugated donkey anti-mouse IgG (Jackson ImmunoResearch, 1:250) and/or 594 nm conjugated goat anti-rabbit IgG (Thermo Fisher Scientific, 1:250). Sections were visualized using a Leica DM 2500 light and fluorescent microscope and photographed with Leica DFC7000T camera, using Leica Application Suite X software.