For histological and immunohistochemical analysis, Achilles tendons with calcaneus were separated and fixed with 10% neutral buffered formalin for 48 hours, then put in 10% ethylenediaminetetraacetic acid (EDTA; pH 7.6) to decalcify for 2 weeks, dehydrated with graded concentrations of ethanol and paraffin embedding. The blocks were sectioned with 4-μm thick slices longitudinally using a Paraffin Microtome and processed for hematoxylin and eosin (H&E) staining, Safranin O and fast green (SOFG, Solarbio, Beijing, China) staining, osteocalcin (Ocn), and p-Smad2/3 immunohistochemistry staining as described previously [22 (link)]. We used an optical microscope (Olympus, Japan) for imaging samples. The paraffin mass of each mouse was cut into 3 serial slices, and the number of positive cells was counted in 3 random visual fields. Ocn antibody, p-Smad2/3 antibody, and other related antibodies were all purchased from Abcam company.
P smad2 3 antibody
Manufactured by Abcam
Sourced in United States
The P-Smad2/3 antibody is a tool used in research laboratories to detect the phosphorylated forms of Smad2 and Smad3 proteins, which are key mediators in the transforming growth factor-beta (TGF-β) signaling pathway. This antibody can be used in various techniques, such as Western blotting, immunohistochemistry, and immunocytochemistry, to analyze the activation and regulation of the TGF-β signaling pathway in different cell and tissue samples.
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2 protocols using p smad2 3 antibody
1
Histological Analysis of Achilles Tendon Decalcification
2
Immunohistochemical Evaluation of TGF-β/SMAD Signaling
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To evaluate TGF-β/SMAD signalling pathways, immunohistochemistry was performed on resin embedded skin sections. After treatment in a saturated solution of sodium hydrate in absolute alcohol for 30 min, the sections were incubated at 37 °C for 1 h with p-SMAD1/5/8 antibody (Millipore Corporation, Oakville, CA, USA) and p-SMAD2/3 antibody (Abcam, Cambridge, UK) at 1:50 dilution using the detection kit ultraview DAB (Roche, Monza, Italy) in an automated staining system BenchMark XT (Roche, Monza, Italy), according to the manufacturer’s instructions. Finally, the slides were counterstained with haematoxylin, mounted, and observed with a Zeiss light microscope.
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