Anti pro il 1β

Manufactured by Santa Cruz Biotechnology

Sourced in United States

Anti-pro-IL-1β is a laboratory reagent used for the detection and quantification of pro-interleukin-1β (pro-IL-1β) in biological samples. It serves as a tool for researchers studying the expression and regulation of this cytokine precursor.

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IL-1β (163 citations) Anti-IL-1β (78 citations) Pro-IL-1β (3 citations)

3 protocols using anti pro il 1β

Quantification of HMGB1 Release in Macrophages

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Replicate wells of THP-1 derived macrophages (4 x 10⁶ cells/well) were incubated as described with Cdt. The cells were washed and treated with 20 mM Tris-HCl buffer (pH 7.5) containing 150 mM NaCl, 1 mM EDTA, 1% NP-40, 1% sodium deoxycholate and protease and phosphatase inhibitors (ThermoFisher Pierce Protein Biology, Pittsburgh, PA); replicate wells were pooled and protein concentration determined. HMGB1 release into culture supernatants was determined following precipitation of supernatants in 20% TCA. Samples were separated on 12% SDS-PAGE and then transferred to nitrocellulose. The membrane was blocked with BLOTTO and then incubated with anti-pro-IL-1β, anti-caspase-4, anti-Syk, anti-pSyk, anti-GSDMD or anti-GAPDH antibody (Santa Cruz Biotechnology) for 18 hr at 4°C (Shenker et al., 1999 (link)). Membranes were washed, incubated with either goat anti-rabbit IgG or goat-anti-mouse IgG conjugated to horseradish peroxidase. The Western blots were developed using chemiluminescence and analyzed (Licor).

Shenker B.J., Walker L.M., Zekavat Z., Ojcius D.M., Huang P.R, & Boesze-Battaglia K. (2020). Cytolethal distending toxin-induced release of interleukin-1β by human macrophages is dependent upon activation of glycogen synthase kinase (GSK) 3β, spleen tyrosine kinase (Syk) and the noncanonical inflammasome. Cellular microbiology, 22(7), e13194.

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Inflammatory Signaling Pathway Analysis

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Ficoll-Paque was purchased from GE Healthcare (Little Halfont, Buckinghamshire, UK). Ultrapure LPS was purchased from InvivoGen (California, USA). Anti-IL-1β, anti-p65, anti-phospho-p65, anti-p38, anti-phospho-p38, anti-ERK, anti-phospho-ERK, anti-JNK, anti-phospho-JNK, anti-c-Jun, anti-phospho-c-Jun, anti-phospho-c-Fos and anti-c-Fos were all purchased from Cell Signaling Technology (Beverly, MA, USA). Anti-G6PD was purchased from Genesis Biotech (Taiwan). ATP, nigericin, PMA, hydrogen peroxide, DPI, NADPH, and anti-β-Actin were purchased from Sigma-Aldrich (St. Louis, MO, USA). Anti-NLRP3 was purchased from AdipoGen Life Sciences (San Diego, CA, USA), and anti-ASC was purchased from Medical & Biological Laboratories (Japan). The anti-caspase-1, anti-pro-IL-1β, anti-mouse and anti-rabbit antibodies were purchased from Santa Cruz Biotechnology (Dallas, TX, USA). Anti-α-tubulin was purchased from Merck Millipore (Burlington, USA). Anti-lamin B1 was purchased from Proteintech (Illinois, USA). Polyvinylidene difluoride (PVDF) membranes and Immobilon Western Chemiluminescent HRP Substrate (ECL) were purchased from Millipore Corporation (Billerica, MA, USA).

Yen W.C., Wu Y.H., Wu C.C., Lin H.R., Stern A., Chen S.H., Shu J.C, & Tsun-Yee Chiu D. (2019). Impaired inflammasome activation and bacterial clearance in G6PD deficiency due to defective NOX/p38 MAPK/AP-1 redox signaling. Redox Biology, 28, 101363.

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Western Blot Analysis of Inflammasome Activation

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Cells derived from 10 NORM and 10 CKD-HD were lysed in RIPA buffer (1 mM phenylmethylsulphonylfluoride, 5 mM EDTA, 1 mM sodium orthovanadate, 150 mM sodium chloride, 8 μg/ml leupeptin, 1.5% NonidetP-40, 20 mM Tris–HCl, pH 7.4). Aliquots containing 45 μg of proteins from each lysate were subjected to SDS–PAGE on a Criterion Tris-HCl 4–20% precast gels and then transferred onto PVDF membrane (Millipore). Membranes were incubated with primary antibodies as follows: anti-cleaved-Caspase-1 (Cell Signaling), anti-proCaspase-1 (Cell Signaling), anti-cleaved-IL1β (Cell Signaling), anti-pro-IL-1β (Santa Cruz), anti-IL-18 (Santa Cruz), and anti-actin (Santa Cruz). Blots were subsequently incubated with secondary antibodies HRP-labelled (Santa Cruz Biotechnology Santa Cruz, CA). Proteins were detected by Chemiluminescence (Amersham, GE Healthcare). Images were acquired using a scanner EPSON Perfection 2580 Photo (EPSON, Long Beach, CA, USA) and quantified by Image J 1.34 Software (http://rsb.info.nih.gov/ij/). The intensity of bands of interest was normalized to the signal intensity of the corresponding procaspase-1, pro-IL-1β, or pro-IL-18 band present on the same membrane. Actin was used as loading control.

Granata S., Masola V., Zoratti E., Scupoli M.T., Baruzzi A., Messa M., Sallustio F., Gesualdo L., Lupo A, & Zaza G. (2015). NLRP3 Inflammasome Activation in Dialyzed Chronic Kidney Disease Patients. PLoS ONE, 10(3), e0122272.

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