Anti NLRP3  

Immunohistochemistry analysis of NLRP3 using anti-NLRP3 antibody. NLRP3 was detected in a paraffin-embedded section of human tonsil tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2 μg/ml rabbit anti-NLRP3 Antibody overnight at 4°C. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using HRP Conjugated Rabbit IgG Super Vision Assay Kit with DAB as the chromogen.

Immunohistochemistry analysis of NLRP3 using anti-NLRP3 antibody. NLRP3 was detected in a paraffin-embedded section of human squamous cell cervical carcinoma tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2 μg/ml rabbit anti-NLRP3 Antibody overnight at 4°C. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using HRP Conjugated Rabbit IgG Super Vision Assay Kit with DAB as the chromogen.

Western blot analysis of NLRP3 using anti-NLRP3 antibody. Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 30μg of sample under reducing conditions. Lane 1: human THP-1 whole cell lysates, Lane 2: rat PC-12 whole cell lysates, Lane 3: mouse spleen whole cell lysates, Lane 4: mouse RAW264.7 whole cell lysates. After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. Blocked the membrane with 5% non-fat milk/TBS for 1.5 hour at RT. The membrane was incubated with affinity purified rabbit anti-NLRP3 antigen polyclonal antibody at 0.5 μg/mL overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:5000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit with Tanon 5200 system. A specific band was detected for NLRP3 at approximately 110 kDa. The expected band size for NLRP3 is at 118 kDa.

Flow Cytometry analysis of THP-1 cells using anti-NLRP3 antibody. Overlay histogram showing THP-1 cells stained with (NLRP3, Blue line). To facilitate intracellular staining, cells were fixed with 4% paraformaldehyde and permeabilized with permeabilization buffer. The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-NLRP3 Antibody (p/n 600-401-R14, 1 μg/1x106？cells) for 30 min at 20°C. DyLightR488 conjugated goat anti-rabbit IgG (5-10 μg/1x106？cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control rabbit IgG antibody (1 μg/1x106, Green line) used under the same conditions. Unlabelled sample (Red line) was also used as a control.