Smad2 Antibody (YZ-13): sc-101153

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	-Mouse (1:200) -Use clean forceps to gently handle the blot from the corner without creasing the membrane. Do not write on the blot with pen or marker, as the ink can fluoresce and cause background.

Protocol tips
-Mouse (1:200) -Use clean forceps to gently handle the blot from the corner without creasing the membrane. Do not write on the blot with pen or marker, as the ink can fluoresce and cause background.

Publication protocol

Total protein was extracted from the rat lung tissues and A549 cells using a total protein extraction kit (cat. no. E211-02; Vazyme, Inc., Nanjing, China). Protein concentrations were determined using a bicinchoninic acid kit (cat. no. P0010; Beyotime Institute of Biotechnology, Haimen, China). Protein samples (50 μg) were separated by 10% SDS-PAGE and then transferred onto a nitrocellulose membrane (EMD Millipore Corp., Billerica, MA, USA). The nitrocellulose membrane was blocked with 3% bovine serum albumin (Amresco Inc., Solon, OH, USA) in Tris-buffered saline overnight at 4°C. After washing with phosphate-buffered saline, the membrane was incubated with the following primary antibodies: rabbit polyclonal anti-E-cadherin (cat. no. sc-7870; 1:800), mouse monoclonal anti-vimentin (cat. no. sc-373717; 1:400), rabbit polyclonal anti-TGF-β1 (cat. no. sc-146; 1:400), mouse monoclonal anti-Smad2 (cat. no. sc-101153; 1:200), all from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA), mouse monoclonal anti-α-smooth muscle actin (cat. no. BM0002; 1:500) or rabbit polyclonal anti-GAPDH (cat. no. BA2913; 1:2,000), both from Boster Biological Technology Ltd., (Wuhan, China), overnight at 4°C. The membrane was then incubated with the horseradish peroxidase (HRP)-labeled secondary antibody [goat anti-mouse immunoglobulin (Ig)G-HRP; sc-2005; 1:4,000) or goat anti-rabbit IgG-HRP (cat. no. sc-2004; 1:4,000; Santa Cruz Biotechnology, Inc.) for 2 h at room temperature. The immunoreactive proteins were detected using an enhanced chemiluminescence kit (cat. no. 32209; Pierce Biotechnolgy, Inc., Rockford, IL, USA). GAPDH was used as a loading control. Images of the blots were captured using a ScanJet 4C Flatbed Scanner (Hewlett-Packard, Palo Alto, CA, USA). Densitometric quantification was performed using Image-Pro Plus software (version 6.0; Media Cybernetics, Inc., Rockville, MD, USA).

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Manufacturer protocol

Download the product protocol from Santa Cruz Biotechnology for Smad2 Antibody (YZ-13): sc-101153 below.

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