The indicated cells were fixed with PFA for 20 min and permeabilized with 0.15% Triton X-100 for 12 min. Then, the cells were blocked with PBS containing 10% FBS for 30 min. For MyHC and Desmin staining, cells were incubated with anti-MyHC Ab (MAB4470, R&D Systems; 1:50) or Desmin (AF5334, Affinity Biosciences, Cincinnati, OH, USA; 1:100) overnight at 4°C in PBS, and then treated with fluorescent secondary Ab (1:200) for 2 h at room temperature in the dark. After incubation with the secondary Ab, cells were stained with 4′,6-diamidino-2-phenylindole (DAPI; Beyotime Biotechnology; 1 μg/ml) for 10 min, and imaged with a confocal microscope (Nikon).
Af5334
Manufactured by Affinity Biosciences
Sourced in United States, United Kingdom
AF5334 is a laboratory instrument designed for performing various analytical techniques. It is a versatile equipment that can be utilized in a range of applications within the biosciences field. The core function of AF5334 is to facilitate data acquisition and analysis.
Automatically generated - may contain errors
Lab products found in correlation
Mab4470, by R&D Systems (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Beyotime (1 mentions)
Ab50339, by Abcam (1 mentions)
Lysate buffer, by Beyotime (1 mentions)
Ab58968, by Abcam (1 mentions)
Polyvinylidene fluoride membrane, by Merck Group (1 mentions)
Enhanced chemiluminescence reagent, by Biosharp (1 mentions)
Df7967, by Affinity Biosciences (1 mentions)
A21020, by Abbkine (1 mentions)
2 protocols using af5334
1
Immunofluorescence Staining of Myogenic Cells
2
Protein Expression Analysis in Podocytes
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Proteins were extracted with lysate buffer (Beyotime, Nantong, China), and the concentration of the extracted proteins was quantified using the bicinchoninic acid method. The loading solution was established such that the protein concentration and volume of the loaded samples were similar. The proteins were separated by electrophoresis, transferred onto a polyvinylidene fluoride membrane (Millipore, Billerica, MA) activated with methanol, blocked with 5% skim milk for 2 h, and incubated with primary antibodies against nephrin (ab58968, Abcam, Cambridge, UK), podocin (ab50339, Abcam, Cambridge, UK), desmin (AF5334, Affinity, Changzhou, China), and β-tubulin (DF7967, Affinity, Changzhou, China) at 4 °C overnight. The following day, the membrane was rinsed and incubated with a secondary antibody labeled with horseradish peroxidase (goat anti-rabbit IgG; A21020, Abbkine, Wuhan, China) for 2 h. After rinsing again, the membrane was exposed to enhanced chemiluminescence reagents (Biosharp, Shanghai, China), and the grayscale values of the bands were determined using ImageJ software (Bethesda, MD).
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!