Whole cell lysates and western blotting were performed as described previously 9 (link). Antibodies against CD105 (ab169545, 1:1000), Smad2 (ab40855, 1:2000), pSmad3 (ab52903, 1:1000) were purchased from Abcam. An antibody against BMP9 (sc514211, 1:500) was purchased from Santa Cruz Biotechnology (USA). Antibodies against Smad1 (6944, 1:1000), pSmad1/5/8 (13820, 1:1000), and pSmad2 (3108, 1:1000) were purchased from Cell Signaling Technology. Antibodies against Smad3 (YM3417, 1:2000), GAPDH (YM3029, 1:20000), and β-tubulin (YM3030, 1:5000) were purchased from ImmunoWay Biotechnology.
Ym3030
Manufactured by Immunoway
Sourced in United States
The YM3030 is a precision laboratory centrifuge designed for a variety of applications. It features a maximum speed of 30,000 RPM and a maximum RCF of 50,000 x g, making it suitable for a wide range of sample processing tasks. The centrifuge is equipped with a brushless motor and a robust construction for reliable and consistent performance.
Automatically generated - may contain errors
Lab products found in correlation
Ym3029, by Immunoway (3 mentions)
Sc514211, by Santa Cruz Biotechnology (1 mentions)
Ab52903, by Abcam (1 mentions)
Ab169545, by Abcam (1 mentions)
Ab40855, by Abcam (1 mentions)
Scanning densitometry, by Bio-Rad (1 mentions)
Sa00001 15, by Proteintech (1 mentions)
Nitrocellulose membrane, by Merck Group (1 mentions)
Ym3403, by Immunoway (1 mentions)
Whatman, by Cytiva (1 mentions)
Ab76228, by Abcam (1 mentions)
E1g1j, by Cell Signaling Technology (1 mentions)
Yt5743, by Immunoway (1 mentions)
Cleaved gsdmd, by Cell Signaling Technology (1 mentions)
D3a3z, by Cell Signaling Technology (1 mentions)
D3r6y, by Cell Signaling Technology (1 mentions)
Cleaved caspase 3, by Cell Signaling Technology (1 mentions)
Horseradish peroxidase hrp conjugated goat anti rabbit secondary antibody, by Cell Signaling Technology (1 mentions)
Ab32503, by Abcam (1 mentions)
Ab59348, by Abcam (1 mentions)
6 protocols using ym3030
1
Western Blot Analysis of TGF-β Signaling
2
Western Blot Analysis of Protein Expression
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Western blotting analysis was used to analyze the expression of proteins. Briefly, cell lysis was performed using radio immunoprecipitation assay lysis buffer containing protease and phosphatase inhibitor cocktails, and the protein concentrations were determined using a BCA assay. Subsequently, proteins (50 μg) were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis for separation, after which they were electro-transferred onto a polyvinylidene fluoride membrane. After blocking with 5% BSA for 3 h at room temperature, the membranes were incubated with primary antibodies (CXCL12, Proteintech, 17402-1-AP; β-tubulin, Immunoway, YM3030; GAPDH, Immunoway, YM3029) at 4°C overnight followed by incubation with secondary antibodies (HRP-conjugated Affinipure Goat Anti-Rat IgG (H + L), Proteintech, and SA00001-15) for 2 h. Immunoreactive bands were visualized using a chemiluminescence kit, and the density of the bands was determined using scanning densitometry (Bio-Rad, Hercules, CA, USA).
3
Western Blot Analysis of Cardiac Proteins
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total protein samples were extracted from the excised myocardial tissue using a protein extraction kit (Cat No. WB0003; Tiande Yue Co., Beijing, China), according to the manufacturer’s instructions. Protein concentration was then determined using a BCA protein assay kit (Cat No. WB0028; Tiande Yue Co.), according to the manufacturer’s instructions. Protein samples were boiled for 5 min, then fractionated by SDS-PAGE (10–15% polyacrylamide gels) and transferred to a nitrocellulose membrane (Whatman; Sigma-Aldrich Corp., St. Louis, MO, USA). The samples were blocked with milk powder for 2 h at room temperature and then incubated with primary antibodies against Runx2 (YM4192; ImmunoWay Biotechnology, Beijing, China), osteopontin (YM3467; ImmunoWay Biotechnology) and β catenin (YM3403; ImmunoWay Biotechnology) at 4°C overnight. After washing five times with TBST (Cat No. WB0043; Tiande Yue Co.), the membranes were incubated with a secondary goat anti-rabbit polyclonal antibody (S004; Tiande Yue Co.) for 1 h at room temperature. Membranes were then washed six times with TBST. Western blot bands were quantified using Total Lab Quant software, version 1.0 (Nature Gene Corp, Beijing, China) by measuring the band intensity for each group and normalizing to β-tubulin (YM3030; ImmunoWay Biotechnology) as an internal control.
4
Comprehensive Immunoblotting Techniques
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Standard techniques were used for protein quantification, separation, transfer, and blotting in KLE and HEC-1A cells. Primary antibodies immunoblotting antibodies were used: ERRα (1: 500, ab76228, Abcam, London, UK), ERRα (1:500, E1G1J, Cell Signaling Technology, Massachusetts, USA), NLRP3 (1:500; 19,771-1AP, Proteintech, Wuhan, China), HIF-1α (1:500; 20,960-1AP, Proteintech, Wuhan, China), cleaved GSDMD (1:500; #36,425, Cell Signaling Technology, Massachusetts, USA), GSDMD (1:500, E9S1X, Cell Signaling Technology, Massachusetts, USA), GSDME (1:500, 84005S, Cell Signaling Technology, Massachusetts, USA), Caspase1 (22,915–1-AP, Proteintech, Wuhan, China), Caspase-1 (YT5743, Immunoway, USA), Caspase-1 (1:500, D7F10, Cell Signaling Technology, Massachusetts, USA), Caspase-3 (1:500, D3R6Y, Cell Signaling Technology, Massachusetts, USA), IL-18 (1:500,10,663–1-AP, Proteintech, Wuhan, China), cleaved-IL-1β (1:500, D3A3Z, Cell Signaling Technology, Massachusetts, USA), β-actin (1:2000, YM3028l, Immunoway, USA), β-tublin (1:2000, YM3030, Immunoway, USA), and GAPDH (1:2000; YM3029, Immunoway, USA).
5
Western Blot Analysis of Apoptosis Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Frozen testis tissue was homogenized in ice-cold radioimmunoprecipitation assay buffer (pH 7.5) containing protease inhibitor cocktail (Roche, USA). After denaturation at 95°C for 5 min, samples were centrifuged at 12,000 × g for 20 min at 4°C and the supernatant was collected and stored at −20°C. Proteins were separated by 10% sodium dodecyl sulfate–polyacrylamide gel electrophoresis and transferred to a polyvinylidene difluoride membrane that was blocked overnight at 4°C in PBS containing 0.1% Tween 20 (PBS-T) and 5% skim milk, and then probed with rabbit anti-rat antibodies against the following proteins: Bcl-2 (1:1000; ab59348) and Bax (1:1000; ab32503) (both from Abcam, USA); cytochrome C (1:500; sc13156) (Santa Cruz Biotechnology, USA); cleaved caspase-9 (1:1000: #9506) and cleaved caspase-3 (1:500; #9664) (both from Cell Signaling Technology, USA); and β-tubulin (1:5000; YM3030) (Immunoway, USA). After three washes with TBS-T, the membrane was incubated at room temperature for 1 h with horseradish peroxidase (HRP)-conjugated goat anti-rabbit secondary antibody (1:1000; Cell Signaling Technology) in PBS-T with 2% skim milk and washed three times with TBS-T [35 (link), 36 (link)]. Immunoreactivity was visualized by enhanced chemiluminescence. The relative signal intensity of protein bands was analyzed with Image J software (National Institutes of Health, USA).
6
Skin Protein Expression Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The protein levels of p63, β1-integrin, Bmp2, Bmp4 and Collagen IV in the skin tissues were analysed by Western blotting. Skin samples from the three groups were lysed using RIPA buffer and centrifuged for 10 min at 15000 g and 4°C to extract total protein. Then, 20 μg of total protein were resolved via SDS-PAGE and transferred to a PVDF membrane, followed by incubation overnight at 4°C with 4 ml of the appropriate dilutions of the following primary rabbit monoclonal antibodies: anti-p63 (1:2000; ab53039; Abcam), anti-β1-integrin (1:5000; ab179471; Abcam), anti-BMP2 (1:1000; ab14933; Abcam), anti-BMP4 (1:2000; ab39973; Abcam), anti-Collagen IV (1:1000; ab6586; Abcam), and anti-β-tubulin (1:10000; YM3030, Immunoway) as an internal control. Then, the membranes were washed three times and incubated at room temperature for 2 h with HRP-conjugated goat anti-rabbit IgG (H+L) at a dilution of 1:10000 (S004, TDY, China). After the membranes were washed three times, the blots were visualized with ECL.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!