To analyse the mRNA expression of angiogenic relative genes in transplanted human ADSCs, total RNA of tissue was extracted and the expression of angiogenesis related human genes was detected by quantitative RT‐PCR as described previously in 2.3 . To measure the proteins extents, the tissues were lysed in Laemmli Sample Buffer (Bio‐Rad) and the total proteins obtained. The proteins extents of HIF‐1α, VEGF, endothelial nitric oxide synthase (eNOS), inducible nitric oxide synthase (iNOS), Collagen III, Elastin and β‐Actin (Cell Signaling Technology) were detected as described previously in 2.4 , and the band intensity was normalized with β‐Actin as the endogenous control.
Collagen 3
Manufactured by Cell Signaling Technology
Sourced in United States
Collagen III is a type of collagen protein found in various connective tissues, including skin, blood vessels, and organs. It plays a crucial role in the structure and function of these tissues.
Automatically generated - may contain errors
Lab products found in correlation
Collagen 1, by Cell Signaling Technology (3 mentions)
β actin, by Cell Signaling Technology (1 mentions)
Laemmli sample buffer, by Bio-Rad (1 mentions)
Tgf β1, by Cell Signaling Technology (1 mentions)
Gsdmd, by Bioss Antibodies (1 mentions)
Caspase 1, by Cell Signaling Technology (1 mentions)
Il 1β, by Cell Signaling Technology (1 mentions)
Bovine serum albumin (bsa), by Cell Signaling Technology (1 mentions)
Radioimmunoprecipitation assay lysis buffer, by CoWin Biotech (1 mentions)
Proliferating cell nuclear antigen (pcna), by Merck Group (1 mentions)
Aurora b, by Abcam (1 mentions)
Aurora b, by Merck Group (1 mentions)
Proliferating cell nuclear antigen (pcna), by Abcam (1 mentions)
Polyvinylidene fluoride membrane, by Merck Group (1 mentions)
Gapdh, by Proteintech (1 mentions)
Gapdh, by Merck Group (1 mentions)
4 protocols using collagen 3
1
Angiogenic Gene Expression in Transplanted ADSCs
2
Cardiac Protein Expression Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total protein was extracted from the heart tissues and H9C2 cells. After protein concentration measurement, these proteins were separated by SDS-PAGE and subsequently transferred to PVDF membranes. Then, the filter membranes were incubated overnight at 4 °C with the primary antibody (Calumenin from Abcam, Collagen I and Collagen III from Cell Signaling). The membranes were further incubated with corresponding secondary antibody for two hours at room temperature and detected using the enhanced chemiluminescence (ECL) system. β-Actin was used as a loading control. The bands were analyzed by the quantity one.
3
Immunohistochemical Analysis of Inflammation Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Tissue sections were removed oxidase and then subjected to antigen repair. The sections were blocked for 30 min at room temperature using 5% bovine serum albumin followed by incubation with the following primary antibodies at 4°C overnight: caspase-1 (1:200) (Cell Signaling Technology,MA, United States), IL-1β (1:100) (Cell Signaling Technology,MA, United States), TGF-β1 (1:200) (Cell Signaling Technology, MA, United States), GSDMD (1:100) (Bioss, Beijing, China), collagen I (1:200) (Cell Signaling Technology, MA, United States) and collagen III (1:200) (Cell Signaling Technology, MA, United States). Next day, the samples were then washed three times for 5 min each using PBST, and the corresponding secondary antibodies were added followed by incubation for 1 h at room temperature. The sections were then stained with diaminobenzidine, and the nuclei were stained with hematoxylin. The sections were then sealed using neutral gum. A fluorescence microscope under white light conditions (Nikon 80i, Japan) was used for imaging.
4
Protein Expression Analysis in Skin Tissues
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
After homogenization, all skin tissues were lysed for 30 min with ice-cold radioimmunoprecipitation assay lysis buffer (CoWin Biosciences, Taizhou, China). Protein lysate (15 μg; concentration determined by a BCA assay) was subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE; CoWin Biosciences, Taizhou, China) on a 6% gel for collagen I and collagen III; 10% gel for Aurora B, CK14, and CK15; 12% gel for PCNA, pH3, and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) at 80 V for 1.5 h, and then transferred to polyvinylidene fluoride membranes (Merck, Darmstadt, Germany). The membranes were blocked with 5% bovine serum albumin in PBS at room temperature for 1.5 h, followed by incubation with primary antibodies against collagen I (1:1000; Cell Signaling Technology, Danvers, MA, United States), collagen III (1:1000; Cell Signaling Technology, Danvers, MA, United States), PCNA (1:1000; Abcam), Aurora B (1:1000; Abcam), pH3 (1:200; Abcam), CK14 (1:1000; Proteintech), CK15 (1:2000; Proteintech), and GAPDH (1:20,000; Proteintech) at 4°C overnight. Quantitative analysis was performed on the immunoreactive bands using ImageJ software (v.1.53k; NIH, Bethesda, MD, United States).
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!