Immunoblotting Analysis of Cardiac Proteins

Protein extracts were prepared from cells and human left ventricular tissue using M-PER and T-PER lysis buffer (Thermo Scientific, NY) respectively [24 (link)]. Immunoblotting was performed using primary antibody specific for CTGF (Santa Cruz, CA), phospho-ERK1/2, Total-ERK1/2 (Cell Signaling, MA) and GAPDH (Ambion, IL) followed by incubation with infrared dye (IRDye)-conjugated secondary antibodies (LI-COR, NE) and immunoreactive bands were visualized under Odyssey scanner (LI-COR, NE). The bands were quantitated using Image Studio Ver3.1 (LI-COR, NE) and normalized by GAPDH.

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Chatterjee A., Barnard J., Moravec C., Desnoyer R., Tirupula K, & Karnik S.S. (2017). Connective tissue growth factor dependent collagen gene expression induced by MAS agonist AR234960 in human cardiac fibroblasts. PLoS ONE, 12(12), e0190217.

Publication 2017

M-PER T-PER lysis buffer phospho-ERK1/2 Total-ERK1/2 GAPDH Odyssey scanner Image Studio Ver3.1

Antibodies Antibody specific Buffer Cells Ctgf Erk1 Gapdh Human Left ventricular Protein Tissue

Corresponding Organization : Bose Institute

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Variable analysis

independent variables

Cell and human left ventricular tissue extracts

dependent variables

Levels of CTGF, phospho-ERK1/2, and Total-ERK1/2 proteins

control variables

GAPDH (used for normalization)

controls

Positive control: None mentioned
Negative control: None mentioned

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