Supernatants from the basolateral side of ALI culture were collected to measure the secretion level of cytokines. IL-6, IL-8, MCP-1, and IFN-β ELISA kits were purchased from R&D Systems. Assays were performed according to manufacturer’s instructions. Absorbance at 450 nm was measured using a microplate spectrophotometer.
Ifn β elisa kit
Manufactured by R&D Systems
Sourced in United States
The IFN-β ELISA kit is a laboratory instrument used for the quantitative determination of interferon beta (IFN-β) levels in biological samples. It provides a sensitive and specific method for measuring IFN-β concentrations.
Automatically generated - may contain errors
Lab products found in correlation
Interleukin 6 (il 6), by R&D Systems (1 mentions)
Mcp 1, by R&D Systems (1 mentions)
β actin, by Merck Group (1 mentions)
Enhanced chemiluminescence, by Bio-Rad (1 mentions)
Adenosine triphosphate (atp), by Merck Group (1 mentions)
Lipopolysaccharides (lps), by Merck Group (1 mentions)
Nigericin, by Merck Group (1 mentions)
Anti iκb, by Cell Signaling Technology (1 mentions)
Poly da dt, by Merck Group (1 mentions)
Anti mouse il 1β ab, by R&D Systems (1 mentions)
Anti phospho iκb, by Cell Signaling Technology (1 mentions)
Bio plex pro mouse cytokine 23 plex assay, by Bio-Rad (1 mentions)
Mda detection kit, by Nanjing Jiancheng (1 mentions)
Lipoxygenase inhibitor screening assay kit, by Cayman Chemical (1 mentions)
6 protocols using ifn β elisa kit
1
Cytokine Secretion Quantification in ALI
2
Protein Expression Analysis by Immunoblotting and ELISA
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To analyze protein expression, cells were harvested and solubilized in disruption buffer containing 50 mM Tris-HCl (pH 7.0), 5% 2-mercaptoethanol, 2% SDS, and 2.75% sucrose. Samples were then subjected to electrophoresis on denaturing polyacrylamide gels, transferred to nitrocellulose membranes, blocked with 5% nonfat milk, and reacted with antibodies against β-actin (Sigma), IRF3 (Santa Cruz Biotechnology), phosphorylated IRF3 (pSer396) (Cell Signaling Technology, Inc.), IκBα and phosphorylated IκBα (Santa Cruz Biotechnology), p65/RelA and phosphorylated p65/RelA (pSer536) (Cell Signaling Technology, Inc.). The membranes were rinsed in phosphate-buffered saline and reacted with donkey anti-rabbit immunoglobulin conjugated to horseradish peroxidase. Protein bands were detected by enhanced chemiluminescence (Bio-Rad). To perform enzyme-linked immunosorbent assays (ELISA), supernatants of cell culture were collected and analyzed using mouse tumor necrosis factor alpha (TNF-α) and IFN-β ELISA kits (R&D systems), and mouse interleukin-6 (IL-6) and IL-1β ELISA kits (eBioscience) according to the manufacturer’s instructions.
3
Molecular Mechanisms of NLRP3 Inflammasome Activation
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LPS, ATP, nigericin and poly dA:dT were purchased from Sigma-Aldrich. Mouse IL-1β, IL-6 and IFN-β ELISA kits were obtained from R&D Systems or Biolegend. Anti-mouse caspase-1 and anti-NLRP3 Abs were obtained from AdipoGen Life Sciences. Anti-mouse IL-1β Ab was obtained from R&D Systems. Anti-apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) and anti-mouse β-actin Abs were purchased from Santa Cruz Biotechnology. Anti-IκB, anti-phospho-IκB and anti-IL-6 Abs were obtained from Cell Signaling Technology.
4
Cytokine Profiling of BMDC Cultures
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Supernatants from cultures of BMDCs cultured with or without ADJ (1%) were collected at 24 hours. Cytokines in cell culture supernatants were quantified using Bio-Plex Pro Mouse Cytokine 23-plex Assay (Bio-Rad, M60009RDPD), IFN-β ELISA kit (R&D Systems, DY8234-05), and IL-18 ELISA (Thermo Fisher Scientific, BMS618-3) according to manufacturer’s protocol.
5
Biomarkers of Oxidative Stress and Inflammation
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Malondialdehyde (MDA) content, interferon-β (IFN-β) level, and ALOX15 activity were determined using commercially available kits according to the manufacturer’s instructions. The MDA detection kit was purchased from Nanjing Jiancheng Bioengineering Institute (Nanjing, China). IFN-β ELISA kit was purchased from the R&D system (Minneapolis, MN, USA). ALOX15 inhibition activity was measured using a Lipoxygenase Inhibitor Screening Assay Kit (Cayman Chemical, Ann Arbor, MI, USA).
6
Quantification of CXCL10 and IFNλ1/β in Samples
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CXCL10 levels in cell-free residual NP swab samples or basolateral media of organoid cultures were quantified using a solid-phase sandwich ELISA (catalog no. DY266; R&D Systems). Briefly, frozen viral transport medium from residual NP swab samples or basolateral medium from organoid cultures was thawed on ice and centrifuged to remove cell debris. ELISA was performed according to the manufacturer’s instructions. IFNλ1 protein concentration in culture basolateral media was measured using a commercial IFNλ1 ELISA kit (catalog no. DY7246; R&D Systems) or IFNβ ELISA kit (DY814-05; R&D Systems). Basolateral media from mock-infected and infected cultures was collected from the bottom chamber of the ALI culture, UV inactivated, and stored at −80°C. The ELISA assay was performed on neat and diluted media per the manufacturer’s instructions.
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