Western Blotting Protein Quantification

Western blotting was performed as described in our previous publications (Lan et al. 2013 (link), 2016 (link); Wen et al. 2019 (link)). The acquired images were analyzed using the NIH image program in the public domain. Primary antibodies used were rabbit anti-Grem1 (Novus Biologicals, NBP1-31150, 1:1000), rabbit anti-P-Smad2/3 (Abcam, ab63399, 1:1000), rabbit anti-P-Smad1/5/8 (Millipore Sigma, AB3848-I, 1:1000), and mouse anti-actin (Santa Cruz Biotechnology, sc-8432, 1:3000). For protein expression quantification, films were scanned using a Canon Scan 9950F scanner.

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Chen J., Xiao H., Xue R., Kumar V., Aslam R., Mehdi S.F., Luo H., Malhotra A., Lan X, & Singhal P. (2023). Nicotine exacerbates diabetic nephropathy through upregulation of Grem1 expression. Molecular Medicine, 29, 92.

Publication 2023

NBP1-31150 sc-8432

Actin Antibodies Biologicals Grem1 Mouse Novus Protein Public domain Rabbit Scan Smad2

Corresponding Organization :

Other organizations : Hangzhou Seventh Peoples Hospital, Zhejiang University, Southwest Medical University, Feinstein Institute for Medical Research, Donald & Barbara Zucker School of Medicine at Hofstra/Northwell

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Variable analysis

independent variables

Protein samples for Western blotting from previous publications (Lan et al. 2013, 2016; Wen et al. 2019)

dependent variables

Protein expression levels of Grem1, P-Smad2/3, and P-Smad1/5/8 as measured by Western blotting

control variables

Experimental procedures for Western blotting as described in previous publications (Lan et al. 2013, 2016; Wen et al. 2019)
Primary antibodies used: rabbit anti-Grem1, rabbit anti-P-Smad2/3, rabbit anti-P-Smad1/5/8, and mouse anti-actin
Image acquisition and analysis using NIH image program
Film scanning using a Canon Scan 9950F scanner

controls

Positive control: No positive control was explicitly mentioned.
Negative control: No negative control was explicitly mentioned.

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