Mouse aortic smooth muscle cells were lysed in Pierce IP Lysis Buffer (87787, Pierce, Waltham, Mass). Floating cells were collected via centrifugation (2000 rpm for 5 minutes) and combined with attached cells then co-immunoprecipitation experiments were performed using SureBeads magnetic beads (1614013, Biorad, Hercules, Calif) according to the manufacturer's protocol. In brief, magnetic beads were washed in PBS saline/Tween (PBST) then incubated with anti-RIPK3 antibody (2283, ProSci, Poway, Calif) or its isotype control for 30 minutes at room temperature. Beads were magnetized and washed 3 times with PBST, then incubated with cell lysate for 1 hour at room temperature. After incubation, beads were washed three times with PBST, and immunoprecipitated proteins were eluted in a 1× Laemmli buffer and subjected to Western blotting.
Anti ripk3 antibody
Manufactured by ProSci
The Anti-RIPK3 antibody is a research-use-only product designed to recognize and bind to the RIPK3 protein. RIPK3 is a serine/threonine-protein kinase that plays a key role in various cellular processes, including programmed cell death and inflammation. This antibody can be used in techniques such as Western blotting to detect and study the RIPK3 protein.
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Lab products found in correlation
4 protocols using anti ripk3 antibody
1
Immunoprecipitation of RIPK3 in Mouse Aortic Smooth Muscle Cells
2
RIPK3 Interaction Proteomics in Mouse Aortic Smooth Muscle Cells
3
RIPK1 and RIPK3 Phosphorylation Analysis
4
Immunoprecipitation Assay for RIPK3
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The immunoprecipitation assay was performed as described previously34 (link). Primary mouse BMDMs seeded into 10-cm dishes were primed with LPS for 2 h and subjected to HS (43 °C for 30 min). After HS, the cells were incubated at 37 °C for 12 h. After washing with DPBS, cells were lysed in Triton X-100 lysis buffer (20 mM Tris, pH 7.4, 137 mM NaCl, 2 mM EDTA, 1% Triton X-100, 10% glycerol, 1 × complete protease Inhibitor). Supernatant was collected, and 500 μg of cell lysate was incubated with 1 μg of anti-RIPK3 antibody (ProSci, 2283) at 4 °C overnight. The lysate immunoprecipitated with anti-IgG (CST, 3900) served as a negative control. The immune complexes were then purified with 30 μl of protein A magnetic beads (Millipore, LSKMAGA02) at 4 °C for 4 h, then centrifuged and washed with lysis buffer. The immunoprecipitated proteins were further analyzed by western blotting.
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