Chrysin (>98% purity) was obtained from Yuanye (Shanghai, China). Chrysin was dissolved in dimethylsulfoxide (<1‰ DMSO) and freshly diluted in culture media for all in vitro experiments. Monoiodoacetic acid, lipopolysaccharide (LPS), type I collagenase and DMSO were all obtained from Sigma-Aldrich (Sigma, St.Louis, MO, USA). TRIzol, Dulbecco’s Modified Eagle Culture Medium (DMEM), Penicillin-Streptomycin mixture, fetal bovine serum (FBS) were purchased from Gibco (Rockville, USA). The primary antibodies for GAPDH, NLRP3, Caspase-1, Interleukin-18, Interleukin-1β and ASC were all purchased from Abcam (Cambridge, UK). Antibodies for c-Fos were purchased from Servicebio (Wuhan, China). Picro Sirius Red Stain kit and Goat anti-rabbit IgG H&L (HRP) were also supplied by Abcam (Cambridge, UK). In addition, 5×HiScript II qRt SuperMix and 2×ChamQ SYBR qPCR MsterMix (Low ROX Premixed) were obtained from Vazyme (Nanjing, China). The primers were supplied by Sangon Biotech (Shanghai, China). Caspase-1 Activity Assay Kit and Cell-Counting Kit-8 were obtained from Solarbio (Beijing, China). Enzyme-linked immunosorbent assays (ELISA) kits for Interleukin-1β and Interleukin-18 were supplied by Invitrogen (Life Technologies Corp., California, USA). ELISA kits for CGRP and SP were supplied by Jin Yibai (Nanjing, China).
Caspase 1 activity assay kit
Manufactured by Solarbio
Sourced in China
The Caspase-1 Activity Assay Kit is a laboratory tool designed to measure the activity of the Caspase-1 enzyme. Caspase-1 is a key player in the inflammatory response and apoptosis (programmed cell death) pathways. The kit provides the necessary reagents and protocols to quantify Caspase-1 activity in various sample types.
Automatically generated - may contain errors
Lab products found in correlation
Microplate reader, by Molecular Devices (2 mentions)
Microplate reader, by Thermo Fisher Scientific (2 mentions)
Goat anti rabbit igg h l hrp, by Abcam (1 mentions)
Trizol, by Thermo Fisher Scientific (1 mentions)
Picro sirius red stain kit, by Abcam (1 mentions)
Chrysin, by Yuanye Bio-Technology (1 mentions)
Dulbecco s modified eagle culture medium dmem, by Thermo Fisher Scientific (1 mentions)
Penicillin streptomycin mixture, by Thermo Fisher Scientific (1 mentions)
Hiscript 2 q rt supermix, by Vazyme (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (1 mentions)
Gapdh, by Abcam (1 mentions)
Nlrp3, by Abcam (1 mentions)
Cell counting kit 8 (cck8), by Solarbio (1 mentions)
Caspase 1, by Abcam (1 mentions)
Pro caspase 1, by Wanlei (1 mentions)
Pro il 1β, by Wanlei (1 mentions)
Rnaios plus, by Takara Bio (1 mentions)
Primescript rt reagent kit, by Takara Bio (1 mentions)
Tb green premix ex taq 2, by Takara Bio (1 mentions)
10 protocols using caspase 1 activity assay kit
1
Chrysin Modulates Inflammatory Pathways
2
Assessing Inflammasome Activation Markers
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Alanine amiotransferase (ALT), triglyceride (TG) and total cholesterol (TC) assay kits were purchased from Nanjing Jiancheng Biotechnology (Nanjing, Jiangsu, China). RNAios Plus, Pri-meScript™ RT reagent kit, and TB Green® Pre-mix Ex Taq™ II were obtained from Takara Biotechnology (Tokyo, Japan). The Caspase-1 Activity Assay Kit and Hoechst 33342/Propidium Iodide (PI) Double Staining Kit were purchased from Solarbio Science & Technology Co., Ltd. (Beijing, China). All other chemicals used were of analytical grade and commercially available.
The following primary antibodies were used: mouse anti-GAPDH (6C5) antibody from Beyotime (Shanghai, China), mouse anti-ASC/TMS1/PYCARD (F-9) antibody from Santa Cruz (Santa Cruz, CA), mouse anti-beta-actin (2D4H5) antibody from Proteintech (Wuhan, China), rabbit polyclonal antibody against GSDMD from Proteintech (Wuhan, China), and rabbit anti-cleaved N-terminal GSDMD (EPR20829-408) antibody from Abcam (Cambridge, UK). The rabbit polyclonal antibodies against NLRP3, pro caspase-1, caspase-1 p20, pro IL-1β, IL-1β, and IL-18 were obtained from Wanleibio (Shenyang, Liaoning, China).
The following primary antibodies were used: mouse anti-GAPDH (6C5) antibody from Beyotime (Shanghai, China), mouse anti-ASC/TMS1/PYCARD (F-9) antibody from Santa Cruz (Santa Cruz, CA), mouse anti-beta-actin (2D4H5) antibody from Proteintech (Wuhan, China), rabbit polyclonal antibody against GSDMD from Proteintech (Wuhan, China), and rabbit anti-cleaved N-terminal GSDMD (EPR20829-408) antibody from Abcam (Cambridge, UK). The rabbit polyclonal antibodies against NLRP3, pro caspase-1, caspase-1 p20, pro IL-1β, IL-1β, and IL-18 were obtained from Wanleibio (Shenyang, Liaoning, China).
3
Quantifying Caspase-1 Activity in Cells
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The caspase-1 activity was detected with Caspase-1 Activity Assay Kit (Solarbio, Beijing). The specific steps were conducted according to the manufacturer’s instructions: First, 2–10 × 106 cells were lysed in 50–100 μl lysis buffer on ice for 10 min. For tissue samples, 3–10 mg tissues were added to 100 μl lysis and were homogenated with a tissue homogenizer and centrifuged. The supernatant was retained. Protein concentrations were detected using the Bradford method, ensuring that the protein concentration was 1–3 μg/μl. A standard curve was prepared using the pNA standard. Then, the optical density of specimen was read on a microplate reader (Molecular Device) at 405 nm. The percentage of caspase-1 activity changes was calculated by the radio of OD405 of the experimental wells to that of the normal wells.
4
Measuring Caspase-1 Activity Assay
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The activity of caspase-1 was measured by using the Caspase-1 Activity Assay Kit (Solarbio, China) in keeping with the manufacturer's instruction. The specimens from tissues and cells were lysed by the lysis buffer. The contents of total proteins were evaluated by Bradford method and a microplate reader was employed to examine the optical density (OD) values at the wavelength of 405 nm, which was utilized to represent caspase-1 activity according to the previous study [38] (link).
5
Quantifying Caspase-1 Activity in Tissues and Cells
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The activity of caspase‐1 was measured by using the Caspase‐1 Activity Assay Kit (BC3810, Solarbio, China) in keeping with the manufacturer's instruction. The specimens from tissues and cells were lysed by the lysis buffer. The contents of total proteins were evaluated by Bradford method, and a microplate reader was employed to examine the optical density (OD) values at the wavelength of 405 nm, which was utilized to represent caspase‐1 activity according to the previous study.25 (link)
6
Caspase-1 Activity Assay Protocol
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The Caspase-1 Activity Assay Kit (Solarbio, Beijing) was utilized to measure Caspase-1 activity. The following steps were conducted exactly as recommended by the manufacturer: First, 2–10 × 106 cells were lysed for 10 min on ice in 50–100 μL lysis buffer. The tissues (3–10 mg) were added to 100 μL lysis buffer, homogenized with a tissue homogenizer, and centrifuged. The supernatant was retained. The Bradford method was used to determine protein concentrations, assuring that the protein concentration was 1–3 μg/μL. A p-nitroaniline colorimetric reference was used to create a standard curve. The optical density of the specimen was then measured at 405 nm using a microplate reader (Molecular Devices). The percentage of Caspase-1 activity changes was calculated by the ratio of OD405 of the experimental wells to that of the normal wells.
7
Quantifying Caspase-1 Activity Assay
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The activity of caspase-1 was measured by using the Caspase-1 Activity Assay Kit (Solarbio, China) in keeping with the manufacturer’s instruction. The specimens from tissues and cells were lysed by the lysis buffer. The contents of total proteins were evaluated by Bradford method and a microplate reader was employed to examine the optical density (OD) values at the wavelength of 405 nm, which was utilized to represent caspase-1 activity according to the previous study [52 (link)].
8
Caspase-1 Activity Assay Protocol
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The caspase-1 activity was measured using a Caspase1 Activity Assay kit (Beijing solarbio science and technology co., ltd. Beijing, China). According to the manufacturer’s instructions, about 50 μg protein from cells was mixed with synthetic tetrapeptide Ac-YVAD-pNA and incubated at 37 °C for 2 h. A standard curve was prepared using the pNA standard. The absorbance was determined at 405 nm with a 96-well plate reader and the caspase1 activity was normalized for total proteins of cell lysates.
9
Quantifying Caspase-1 Activity Assay
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Caspase-1 activity was detected using the Caspase-1 Activity Assay Kit (Solarbio). The specific steps were performed according to the manufacturer’s instructions (Solarbio). The protein concentration was ensured from 1 μg/μl to 3 μg/μl. A standard curve was prepared using the pNA standard. Then, the optical density of the specimen was read on a microplate reader (Thermo Fisher Scientific) at 405 nm. The percentage of changes in caspase-1 activity was calculated by the ratio of OD405 of the experimental well to that of the normal well.
10
Caspase-1 Activity Assay Protocol
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Caspase‐1 activity was detected by Caspase‐1 Activity Assay Kit (Solarbio Biotech) according to the manufacturer's instructions. Briefly, for cell samples, 2‐10 × 106 cells were lysed in 50‐100 μl lysis buffer on ice for 10 min. For tissue samples, 3‐10 mg tissues were added to 100 μl lysis and were homogenated with a tissue homogenizer. The supernatant from extracts was collected by centrifuged at 12 000 g for 10 min. Protein concentrations were detected using the Bradford method, ensuring that the protein concentration was 1‐3 μg/μl. A standard curve was prepared using the pNA standard. 10‐35 μl volume of supernatant was incubated with 5 μl 2 mM Ac‐YVAD‐pNA substrate at 37°C. Then, the absorbance values of pNA were read on a spectrophotometer at the optical density 405nm (OD405). The changes of caspase‐1 activity were calculated by the radio of OD405 of the experimental wells to that of the control wells.
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