Total protein of the cochleae was isolated with RIPA (Beyotime) lysis buffer supplemented with PMSF (Beyotime). Protein concentrations were measured using a BCA protein assay kit (Beyotime), and proteins were separated on a BeyoGel™ Plus Precast PAGE Gel for Tris-Gly System (Beyotime) and transferred onto polyvinylidene difluoride membranes (Beyotime). The membranes were blocked with 10% nonfat dried milk in Tris-buffered saline with Tween 20 (TBST, 50 mM Tris-HCl (pH 7.4), 150 mM NaCl, and 0.1% Tween 20) for 1 h at room temperature and then blotted overnight with primary antibodies at 4 °C. The primary antibodies were as follows: rabbit anti-Yap (1:1000 dilution) (CST), rabbit anti-phospho-Yap (1:1000 dilution) (Ser127) (CST), rabbit anti-LATS1 (1:1000 dilution) (CST), rabbit anti-phospho-LATS1 (1:1000 dilution) (Thr1079) (CST), rabbit anti-Cyr61 (1:500 dilution) (Santa Cruz), and goat anti-CTGF (1:500 dilution) (Santa Cruz). All blots derive from the same experiment and that they were processed in parallel.
Goat anti ctgf
Manufactured by Santa Cruz Biotechnology
Sourced in United States
Goat anti-CTGF is a polyclonal antibody raised in goats against the Connective Tissue Growth Factor (CTGF) protein. This antibody is designed for use in various immunological techniques to detect and study the CTGF protein.
Automatically generated - may contain errors
Lab products found in correlation
Las 3000 imaging workstation, by Fujifilm (2 mentions)
Horse anti mouse, by Cell Signaling Technology (2 mentions)
Biotinylated secondary antibody, by Vector Laboratories (2 mentions)
Mouse anti neun, by Merck Group (2 mentions)
Rabbit anti cleaved caspase 3, by Cell Signaling Technology (2 mentions)
Fitc isolectin b4, by Vector Laboratories (2 mentions)
Alexa fluor conjugated secondary antibody, by Thermo Fisher Scientific (2 mentions)
Rabbit anti pten, by Abcam (2 mentions)
Mouse anti parvalbumin, by Swant (2 mentions)
Rabbit anti dsred, by Takara Bio (2 mentions)
Rabbit anti gaba, by Merck Group (2 mentions)
Rat anti somatostatin, by Merck Group (2 mentions)
Mouse anti gaba, by Merck Group (2 mentions)
Goat anti mcherry, by Antibodies-online (2 mentions)
Alexa fluor conjugated streptavidin, by Vector Laboratories (2 mentions)
Isolectin b4 dylight 594, by Vector Laboratories (2 mentions)
Rabbit anti parvalbumin, by Swant (2 mentions)
Polyvinylidene difluoride membrane, by Beyotime (1 mentions)
Radioimmunoprecipitation assay (ripa), by Beyotime (1 mentions)
Phenyl methyl sulfonyl fluoride (pmsf), by Beyotime (1 mentions)
7 protocols using goat anti ctgf
1
Western Blot Analysis of Cochlear Proteins
2
Western Blot Analysis of CD44 and CTGF
3
Protein Expression Analysis of Renal Cortex and HUVECs
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Proteins from the renal cortex or HUVECs were extracted in RIPA buffer with proteinase inhibitors, and protein concentrations were determined using the BCA assay. Twenty-five microgram of proteins were separated by SDS-PAGE and transferred onto PVDF membranes. After blocking with Roti-block (Roth, Karlsruhe, Germany) for 1 h at room temperature, the membranes were incubated with primary antibodies overnight at 4°C and corresponding secondary antibodies for 1 h at room temperature. The proteins were visualized using a chemiluminescent peroxidase substrate (Roche, Mannheim, Germany; or Thermo Scientific, Rockford, USA). Protein expression was quantified using Image J (NIH, USA). Specific primary antibodies used: mouse-anti-α-SMA (Sigma-Aldrich, A5228, 1:2,000), goat-anti-CTGF (Santa Cruz, sc-14939, 1: 200), mouse-anti-γ-tubulin (Sigma, T6557, 1:10,000). The secondary antibodies conjugated with horseradish peroxidase: rabbit-anti-goat (Sigma-Aldrich, A8919), horse-anti-mouse (Cell Signaling, 7076, 1:20,000).
4
Western Blot Analysis of Hippo Pathway
5
Perfusion, Fixation, and Immunostaining of Mouse Brains
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Mice were anaesthetised with an overdose of sodium pentobarbital and
transcardially perfused with saline followed by 4% paraformaldehyde (PFA).
Brains from pups younger than P6 were post-fixed for 4 h while brains from mice
older than P6 were post-fixed for 2 h at 4°C. Brains were sectioned
either on the sliding microtome at 30 or 40 µm as previously
described52 (link) or on a vibratome at 40
or 60 µm. All primary and secondary antibodies were diluted in PBS
containing 0.25% Triton X-100 and 2% BSA. The following antibodies were used:
goat anti-CTGF (1:200, Santa Cruz), rabbit anti cleaved-caspase-3 (1:200, Cell
Signalling), rabbit anti-dsRed (1:500, Clontech), goat anti-mCherry (1:500,
Antibodies-online), rabbit anti-GABA (1:2000, Millipore), mouse anti-GABA
(1:500, Sigma), mouse anti-NeuN (1:500, Millipore) mouse anti-parvalbumin
(1:1000, Swant), rabbit anti-parvalbumin (1:5000, Swant), rat anti-somatostatin
(1:300, Millipore) and rabbit anti-PTEN (1:500, Abcam). We used Alexa
Fluor-conjugated secondary antibodies (Invitrogen). For biotin amplification,
sections were incubated with biotinylated secondary antibody (1:200, Vector
labs), followed by Alexa Fluor-conjugated Streptavidin (1:200, Vector labs).
Blood vessels were stained with Isolectin-B4-FITC or Isolectin-B4-Dylight 594
(1:500, Vector labs).
transcardially perfused with saline followed by 4% paraformaldehyde (PFA).
Brains from pups younger than P6 were post-fixed for 4 h while brains from mice
older than P6 were post-fixed for 2 h at 4°C. Brains were sectioned
either on the sliding microtome at 30 or 40 µm as previously
described52 (link) or on a vibratome at 40
or 60 µm. All primary and secondary antibodies were diluted in PBS
containing 0.25% Triton X-100 and 2% BSA. The following antibodies were used:
goat anti-CTGF (1:200, Santa Cruz), rabbit anti cleaved-caspase-3 (1:200, Cell
Signalling), rabbit anti-dsRed (1:500, Clontech), goat anti-mCherry (1:500,
Antibodies-online), rabbit anti-GABA (1:2000, Millipore), mouse anti-GABA
(1:500, Sigma), mouse anti-NeuN (1:500, Millipore) mouse anti-parvalbumin
(1:1000, Swant), rabbit anti-parvalbumin (1:5000, Swant), rat anti-somatostatin
(1:300, Millipore) and rabbit anti-PTEN (1:500, Abcam). We used Alexa
Fluor-conjugated secondary antibodies (Invitrogen). For biotin amplification,
sections were incubated with biotinylated secondary antibody (1:200, Vector
labs), followed by Alexa Fluor-conjugated Streptavidin (1:200, Vector labs).
Blood vessels were stained with Isolectin-B4-FITC or Isolectin-B4-Dylight 594
(1:500, Vector labs).
6
Protein Isolation and Western Blot Analysis
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Proteins were isolated following the RNA isolation according to the manufacturer’s recommendations (Peqlab). Proteins were dissolved in 1% SDS containing protease and phosphatase inhibitors, and protein content was measured with the bicinchoninic acid assay (Interchim, Montluçon Cedex, France). Western blot analysis was performed with specific antibodies as described previously (Fuchshofer et al. 2011 (link), 2007 (link)). Specific antibodies were used as follows: goat anti-CTGF (1:500, Santa Cruz Biotechnology, Santa Cruz, USA), rabbit anti-α-tubulin (1:2500, Rockland Immunochemicals Inc., Gilbertsville, USA), donkey anti-goat (1:2000, Bethyl Laboratories Inc., Montgomery, USA), and goat anti-rabbit (1:5000, Cell Signaling Technology, Danvers, USA). Chemiluminescence was detected on a LAS 3000 imaging workstation (Fujifilm, Düsseldorf, Germany). For normalization of the signal intensity, α-tubulin was used as a loading control. The intensity of the bands detected by Western blotting was determined using appropriate software (AIDA Image analyzer software, Raytest, Straubenhardt, Germany).
7
Perfusion, Fixation, and Immunostaining of Mouse Brains
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