Cells (8×103 cells/well) were seeded in 24-well plates and treated with PA (0.50 mM) for 6 h with or without 3-MA (5 mM) or N-Acetyl-cysteine (NAC; 1 mM, S0077; Beyotime). Subsequently, cells were fixed in 4% paraformaldehyde (P0099; Beyotime) for 15 min at room temperature and washed twice with PBS, then permeabilized with 0.2% triton X-100 (P0096; Beyotime) for 5 min at room temperature. Thereafter, cells were washed with PBS, incubated with 5% BSA for 1 h, and incubated in anti-LC3 antibody (ab51520, 1:200; Abcam) overnight at 4°C. The cells were then incubated with goat anti-rabbit secondary antibody labeled with fluorescein (ZF-0511, 1:400; ZSGB-BIO) for 1 h at room temperature before staining with DAPI (C1006; Beyotime). Samples were photographed using a wide field fluorescent microscope (IX71; Olympus Corporation). The number of LC3 positive cells in each microscopic field was divided by the number of nuclei in the same field, and was regarded as the rate of LC3 positive cells.
S0077
Manufactured by Beyotime
Sourced in China
The S0077 is a laboratory equipment designed for general laboratory use. It serves as a device for performing various experimental procedures that require consistent and controlled temperature conditions. The core function of the S0077 is to maintain a stable temperature environment for samples or materials under investigation.
Automatically generated - may contain errors
Lab products found in correlation
3 protocols using s0077
1
Autophagy Induction and Inhibition Assay
2
Cardiomyocyte hypoxia/reoxygenation model
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Primary cardiomyocytes were separated and maintained in DMEM containing 4.5 g/L of glucose and supplemented with 10% (v/v) FBS, and 1% penicillin/streptomycin at 37°C in a humidified 5% CO2 incubator. The cardiomyocytes were pretreated with 2.5, 5, and 10 µM emodin (E106693; Aladdin, Shanghai, China), 5 µM Bay-117082 (NF-κB pathway inhibitor) (S2913; Selleck, Shanghai, China), 10 µM NLRP3 inflammasome inhibitor (S3680; Selleck), or N-acetylcysteine (NAC, 1 mM, S0077; Beyotime, Shanghai, China) for 1 hour before the cardiomyocytes were stimulated with hypoxia/reoxygenation (H/R).
3
Inflammatory Responses of Aortic Valve Interstitial Cells
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HAVICs were isolated and cultured as previously reported [17 (link)]. In brief, valve leaflets were subjected to digestion with collagenase, and cells were collected by centrifugation. The cells were cultured in M199 growth medium (Gibco, C11150500BT) containing penicillin G, streptomycin, and 10% fetal bovine serum. Cells from passages 3-6 that reached 80-90% confluence were used for this study.
To determine the influence of S18 on the inflammatory responses in LPS-stimulated HAVICs, HAVICs were treated with LPS (200 ng/mL, Sigma, L4391) in the absence or presence of S18 (0.5 μM) for 24 hours.
To evaluate the role of NF-κB in the suppression of HAVIC inflammation by S18, HAVICs were treated with LPS (200 ng/mL) in the absence or presence of S18 (0.5 μM) for 4 hours.
To determine whether mitochondrial stress is involved in the LPS-induced HAVIC inflammatory responses, HAVICs were treated with NAC (1 mM, Beyotime, S0077) for 1 hour prior to LPS stimulation.
To evaluate the role of IL-37 in mediating the effect of S18 on HAVICs, HAVICs were treated with IL-37 siRNA (50 nM, OBiO) for 8 hours followed by stimulation with LPS with or without S18 (0.5 μM).
To determine the role of IL-37 in attenuating the mitochondrial stress in HAVICs, HAVICs were preincubated with recombinant IL-37 (0.1 ng/mL, MCE, HY-P70455) for 1 hour followed by LPS (200 ng/mL) stimulation.
To determine the influence of S18 on the inflammatory responses in LPS-stimulated HAVICs, HAVICs were treated with LPS (200 ng/mL, Sigma, L4391) in the absence or presence of S18 (0.5 μM) for 24 hours.
To evaluate the role of NF-κB in the suppression of HAVIC inflammation by S18, HAVICs were treated with LPS (200 ng/mL) in the absence or presence of S18 (0.5 μM) for 4 hours.
To determine whether mitochondrial stress is involved in the LPS-induced HAVIC inflammatory responses, HAVICs were treated with NAC (1 mM, Beyotime, S0077) for 1 hour prior to LPS stimulation.
To evaluate the role of IL-37 in mediating the effect of S18 on HAVICs, HAVICs were treated with IL-37 siRNA (50 nM, OBiO) for 8 hours followed by stimulation with LPS with or without S18 (0.5 μM).
To determine the role of IL-37 in attenuating the mitochondrial stress in HAVICs, HAVICs were preincubated with recombinant IL-37 (0.1 ng/mL, MCE, HY-P70455) for 1 hour followed by LPS (200 ng/mL) stimulation.
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