At 12 h after HSR, total protein was extracted from the hippocampal tissues (n=6 per group) as described in the IL-1β and IL-18 assay. Samples containing 30 µg protein quantified by bicinchoninic acid (BCA) assay were separated by 10% SDS-PAGE and transferred onto a PVDF membrane. The membrane was blocked with 5% skimmed milk at 37°C for 2 h, and primary antibodies rabbit anti-rat polyclonal cleaved caspase-1 (1:1,000; cat. no. ab1872; Abcam), monoclonal NLRP3 (1:1,000; cat. no. ab210491; Abcam), polyclonal Gasdermin D (GSDMD; 1:1,000; cat. no. ab219800; Abcam), polyclonal IL-1b (1:1,000; cat. no. K107559P; Beijing Solarbio Science & Technology Co., Ltd.) and polyclonal IL-18 (1:1,000; cat. no. K002143P; Beijing Solarbio Science & Technology Co., Ltd.) were applied at 4°C overnight, followed by incubation with a horseradish peroxidase-conjugated goat anti-rabbit IgG secondary antibody (1:2,000; cat. no. sc-2004; Santa Cruz Biotechnology, Inc.) at 25°C for 1 h. Protein bands in each treatment group were detected using an enhanced chemiluminescence western blot detection system ImageJ 1.48u software (National Institutes of Health). β-actin (1:2,000; cat. no. sc-47778; Santa Cruz Biotechnology, Inc.) was used as an internal reference (20 (link)).
Il 18
Manufactured by Solarbio
Sourced in China
IL-18 is a lab equipment product that measures the levels of interleukin-18 (IL-18) in biological samples. IL-18 is a pro-inflammatory cytokine involved in various immune responses.
Automatically generated - may contain errors
Lab products found in correlation
Il 1β, by Solarbio (3 mentions)
Gasdermin d gsdmd, by Abcam (1 mentions)
Il 1b, by Abcam (1 mentions)
Dcfh da probe, by Beyotime (1 mentions)
Cell culture reagents, by Thermo Fisher Scientific (1 mentions)
Cellevent senescence green flow cytometry assay kit, by Thermo Fisher Scientific (1 mentions)
Nigericin, by MedChemExpress (1 mentions)
β actin, by Cell Signaling Technology (1 mentions)
Senescence β galactosidase β gal staining kit, by Beyotime (1 mentions)
Mcc950, by MedChemExpress (1 mentions)
Nlrp3, by Adipogen (1 mentions)
Microplate reader, by Molecular Devices (1 mentions)
Tnf α, by Solarbio (1 mentions)
4 protocols using il 18
1
Hippocampal Protein Analysis after HSR
2
Senescence and NLRP3 Inflammasome Assay
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We obtained the cell culture reagents from Invitrogen (Carlsbad, CA, USA). We purchased D-galactose (D-gal), N-acetyl-L-cysteine (NAC, ROS scavenger) from Sigma-Aldrich (St. Louis, MO, USA), and MCC950 (NLRP3 inflammasome inhibitor), nigericin (NLRP3 inflammasome activator) from MedChem Express (Monmouth Junction, NJ, USA). The senescence β-galactosidase (β-gal) Staining Kit and DCFH-DA probe were purchased from Beyotime (Shanghai, China). The CellEvent™ Senescence Green Flow Cytometry Assay Kit was purchased from Thermo Fisher Scientific (Waltham, MA, USA). Primary antibodies including P53 (1:1000, Cell Signal Technology, Danvers, MA, USA), P21(1:1000, Cell Signal Technology), β-actin(1:1000, Cell Signal Technology), NLRP3 (1:1000, Adipogen, San Diego, CA, USA) and ASC (1:1000, Abcam, UK) were purchased from the above accompanied. The IL-1β, IL-18 and LDH kits were purchased from Solarbio Technology (Beijing, China).
3
Serum Cytokine Profiling by ELISA
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4
ELISA Quantification of Inflammatory Mediators
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After treatment, the culture medium was collected and processed for ELISA. Culture medium (100 µl) was reacted with the following ELISA kits: PGE2 (cat. no. SEKH-0414; Beijing Solarbio Science & Technology Co., Ltd.), TNF-α (cat. no. SEKH-0047; Beijing Solarbio Science & Technology Co., Ltd.), IL-1β (cat. no. SEKH-0002; Beijing Solarbio Science & Technology Co., Ltd.) and IL-18 (cat. no. SEKH-0028; Beijing Solarbio Science & Technology Co., Ltd.) according to the manufacturers' protocols. The absorbance was measured at 450 nm using a microplate reader (Molecular Devices, LLC).
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