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Asc sc 22514 r

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The ASC (sc-22514-R) is a lab equipment product offered by Santa Cruz Biotechnology. It is a protein that plays a central role in the formation of the inflammasome, a protein complex involved in the activation of inflammatory responses. The ASC product provides researchers with a tool to study this important cellular process.

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Lab products found in correlation

Sc 66846, by Santa Cruz Biotechnology (1 mentions) Caspase 1, by Cell Signaling Technology (1 mentions) X ray film, by Kodak (1 mentions) Enhanced chemiluminescence system, by GE Healthcare (1 mentions) F4 80 mca497ga, by Bio-Rad (1 mentions) Il 1β ab9722, by Abcam (1 mentions) Nrf2 sc 13032, by Santa Cruz Biotechnology (1 mentions) Caspase 1, by Santa Cruz Biotechnology (1 mentions) Gapdh, by Santa Cruz Biotechnology (1 mentions) Nlrp3 ag 20b 0014, by Adipogen (1 mentions) Horseradish peroxidase conjugated secondary antibody, by Bio-Rad (1 mentions) β actin, by Merck Group (1 mentions) Propidium iodide, by Thermo Fisher Scientific (1 mentions) Z vad fmk, by R&D Systems (1 mentions) Actin ac 74, by Merck Group (1 mentions) Yvad fmk, by R&D Systems (1 mentions) Lipofectamine 2000, by Thermo Fisher Scientific (1 mentions)

4 protocols using asc sc 22514 r

1

Western Blot Analysis of Inflammatory Markers

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Western Blot studies were carried out as described previously (45 (link)–47 (link)). Briefly, control and experimental cells were harvested, lysed in RIPA buffer containing 50 mM Tris-Cl (pH 7.5), 150 mM NaCl, 1mM EDTA, 1% NP-40, 0.25% Deoxycholate, 0.1% SDS, 1X protease inhibitor cocktail (Calbiochem, Cocktail Set I), 1mM PMSF, and 0.2mM sodium orthovanadate. Protein concentration was determined using the Bio-Rad Protein Assay kit (PIERCE, Rockford, IL). Total protein lysed extracts (30 μg/lane) were loaded on a 10 % polyacrylamide (PAGE) premade gel (Bio-Rad, Hercules, CA) and after transferring onto PVDF membrane were processed for immunostaining with primary antibodies against APOL1 (anti-mouse, #66124-I-IG, Protein tech), NLRP3 (anti-mouse, #sc-66846; Santa Cruz); ASC (sc-22514-R; Santa Cruz Biotechnology), Caspase-1and Cleaved (C ) Caspase-1 (antirabbit #4199;1:700, Cell Signaling): followed by treatment with horseradish peroxidase-labeled appropriate secondary antibodies. The blots were developed using a chemiluminescence detection kit (PIERCE, Rockford, IL) and exposed to X-ray film (Eastman Kodak Co., Rochester, NY). Equal protein loading and the protein transfers were confirmed by immunoblotting for determination of GAPDH protein using a monoclonal GAPDH antibody (#SC-47724; 1:3000, Santa Cruz) performed on the same (stripped) western blots.
Jha A., Kumar V., Haque S., Ayasolla K., Saha S., Lan X., Malhotra A., Saleem M.A., Skorecki K, & Singhal P.C. (2019). Alterations in Plasma Membrane Ion Channel Structures Stimulate NLRP3 Inflammasomes Activation in APOL1 Risk Milieu. The FEBS journal, 287(10), 2000-2022.
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2

Protein expression analysis in kidney

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Kidney and cell lysates were extracted with extraction buffer or sample buffer as described previously43 (link). Protein samples were subjected to immunoblotting analysis with antibodies against α-SMA (100M4795; Sigma-Aldrich, St Louis, MO, USA), ASC (sc-22514-R; Santa Cruz Biotechnology, Dallas, TX, USA), caspase-1 (sc-514; Santa Cruz Biotechnology), F4/80 (MCA497GA; AbD Serotec, Raleigh, NC, USA), IL-1β (ab9722; Abcam, Cambridge, MA, USA), Nrf2 (sc-13032; Santa Cruz Biotechnology) and GAPDH (sc-25778; Santa Cruz Biotechnology). Signals were detected using an enhanced chemiluminescence system (GE Healthcare Japan, Tokyo, Japan). Each western blot was performed on a pooled sample of 4–6 mice from each group.
Sogawa Y., Nagasu H., Iwase S., Ihoriya C., Itano S., Uchida A., Kidokoro K., Taniguchi S., Takahashi M., Satoh M., Sasaki T., Suzuki T., Yamamoto M., Horng T, & Kashihara N. (2017). Infiltration of M1, but not M2, macrophages is impaired after unilateral ureter obstruction in Nrf2-deficient mice. Scientific Reports, 7, 8801.
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3

Western Blot Analysis of Inflammasome Proteins

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Cells were harvested and centrifuged, then they were lysed in loading buffer (62,5mM Tris pH = 8.8, 25% glycerol, 2% SDS, 1% β-mercaptoethanol and 1% BPB). Before loading all samples were boiled for 10 minutes. Proteins were separated by SDS-PAGE and transferred onto nitrocellulose membranes. Membranes were then blocked with 5% non-fat milk, washed briefly, incubated with primary antibodies at 4°C overnight. Pro-IL-1β (AF401-NA) was from R&D System, ASC (sc22514-R) was from Santa Cruz, pro-caspase-1 (AG-20B-0042) and NLRP3 (AG-20B-0014) antibodies were obtained from AdipoGen. Primary antibodies were incubated with corresponding horseradish peroxidase-conjugated secondary antibodies from BioRad for 1 hour at room temperature. Proteins were visualized by Supersignal West-Pico peroxide/luminol enhancer solution from Pierce. To verify the loading of equal amount of protein sample, the β-actin (Sigma-Aldrich) expression was detected.
Czimmerer Z., Daniel B., Horvath A., Rückerl D., Nagy G., Kiss M., Peloquin M., Budai M.M., Cuaranta-Monroy I., Simandi Z., Steiner L., Nagy B J.r., Poliska S., Banko C., Bacso Z., Schulman I.G., Sauer S., Deleuze J.F., Allen J.E., Benko S, & Nagy L. (2018). The Transcription Factor STAT6 Mediates Direct Repression of Inflammatory Enhancers and Limits Activation of Alternatively Polarized Macrophages. Immunity, 48(1), 75-90.e6.
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4

Immunoblotting Antibodies and Inhibitors

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Antibodies for immunoblotting were from Sigma (actin, AC-74 used at 1:5,000 dilution and caspase-11, 17D9 used at 1:1,000 dilution), Cell Signaling (caspase-7, 9492 used at 1:1,000 dilution), Adipogen (caspase-1 p20, AG-20B-0042 used at 1:1,000 dilution), DSHB (tubulin, E7 used at 1:5,000 dilution) and Santa Cruz Biotechnology (ASC sc-22514-R used at 1:1,000 dilution). Propidium iodide was from Life Technologies and Lipofectamine2000 from Invitrogen. siRNAs for caspase-7 and caspase-11 were purchased from Santa Cruz and used at 40 pmol. zVAD-FMK and YVAD-FMK were from R&D systems.
Thurston T.L., Matthews S.A., Jennings E., Alix E., Shao F., Shenoy A.R., Birrell M.A, & Holden D.W. (2016). Growth inhibition of cytosolic Salmonella by caspase-1 and caspase-11 precedes host cell death. Nature Communications, 7, 13292.
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