Rats were decapitated and their ACC were removed. Next, we centrifuged tissue samples at 12,000× g for 20 min at 4 °C and quantified target proteins. Gel electrophoresis (SDS-PAGE) was used to separate proteins, which were then electro-transferred to PVDF membranes (Millipore, Billerica, MA USA). After being sealed with 5% skimmed milk at room temperature for 1 h, the membranes were placed in anti-NLRP3 antibody (1:1000, Affinity, Cincinnati, OH, USA) and anti-Piezo1 antibody (1:1000, Alomone, Jerusalem, Israel) overnight at 4 °C and then incubated with HRP-conjugated donkey anti-mouse, anti-goat, or anti-rabbit secondary antibody (1:10,000, Abcam, Cambridge, UK). β-actin (1:1000, Boster, Wuhan, China) was used as a load control. Enhanced chemiluminescence (Bio-Rad, Hercules, CA, USA) was used to detect the target protein bands, which were imaged using the Tanon-5200 chemiluminescence imaging system (Tanon Science and Technology, Shanghai, China). An image analysis system (KONTRON IBAS 2.0, Germany) was used to quantify the protein level in comparison to the expression level of β-actin.
Anti nlrp3 antibody
Manufactured by Affinity Biosciences
Sourced in United States
The Anti-NLRP3 antibody is a laboratory reagent designed to detect and study the NLRP3 protein. NLRP3 is a key component of the inflammasome, a protein complex involved in the immune response. The antibody can be used in various research applications, such as Western blotting, immunohistochemistry, and immunoprecipitation, to investigate the expression and localization of NLRP3 in biological samples.
Automatically generated - may contain errors
Lab products found in correlation
Pvdf membrane, by Merck Group (1 mentions)
β actin, by Boster Bio (1 mentions)
Enhanced chemiluminescence, by Bio-Rad (1 mentions)
Confocal fluorescence microscope, by Leica (1 mentions)
Anti cleaved caspase 1, by Affinity Biosciences (1 mentions)
Anti vp1, by Merck Group (1 mentions)
Diamidino 2 phenylindole dapi, by Beyotime (1 mentions)
2 protocols using anti nlrp3 antibody
1
Quantification of NLRP3 and Piezo1 Proteins
2
Immunofluorescence Assay for VP1, Caspase-1 and NLRP3
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The pre-cooled PBS was used to rinse the cells in 24-well plates (2 × 104 cells/well) for 3 times and 4% paraformaldehyde was employed to fix the cells deposited in plates of 24-well. After that, cells were subjected to permeabilization with PBS containing 0.1% Triton X-100 for 10 min at 37 °C. Then, 3% BSA was added for blockading the nonspecific binding. Next, the cells were incubated with the anti-VP1 (1:1000 dilution; Millipore, USA), anti-cleaved-Caspase1 (1:100 dilution; Affinity, USA) and anti-NLRP3 antibody (1:100 dilution; Affinity, USA) overnight at 4 °C, followed by incubation with fluorescein isothiocyanate (FITC)-conjugated donkey anti-mouse IgG and Daylight 594-conjugated donkey anti-rabbit IgG secondary antibodies (1:300 dilution; CST, USA) for 1 h. Finally, PBS was adopted to rinse the cells for 3 times and diamidino-2-phenylindole (DAPI; 1:1000 dilution; Beyotime, China) was taken to stain them at 37 °C for 5 min. With the help of a confocal fluorescence microscope (Leica, Germany), the images were obtained.
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