24 Sprague-Dawley (SD) rats (males, 200–220 g) were used for subcutaneous implantation of scaffolds. All animal experimental protocols followed the guidelines and regulations of the Institutional Animal Care and Use Committee of Kunming Medical University (China). The rats were randomly divided into four groups (six rats/group): PU/n-HA, 0.5%, 1%, and 2% gastrodin-PU/n-HA. Each sterilized disc specimen (Ф 13 mm × 2 mm) was symmetrically transplanted into the dorsal subcutaneous pocket of each rat. All rats were then sacrificed at 3 and 14 days after implantation. Collected blood serum was assayed to measure the content of IL-1β using an IL-1β ELISA kit (Nanjing Jiancheng, China) according to the manufacturer's instructions. Vascular connections ex vivo were tested, with further details reported in the SI. Samples with the surrounding tissues along with the heart, liver, spleen, kidney, and lung were collected and fixed with 4% paraformaldehyde solution for 7 days. The tissue samples were dehydrated in graded concentrations of ethanol. After this, they were cleared in xylene, embedded in paraffin wax, and cut into 5-μm thick sections. The sections were deparaffinized, rehydrated, and stained hematoxylin and eosin (H&E), Masson, CD31, Arg-1, and iNOS, followed by imaging using an optical microscope.
Il 1β elisa kit
Manufactured by Nanjing Jiancheng
Sourced in China
The IL-1β ELISA kits are a laboratory tool used to detect and quantify the presence of the inflammatory cytokine interleukin-1 beta (IL-1β) in biological samples. The kits utilize the enzyme-linked immunosorbent assay (ELISA) technique to measure IL-1β levels accurately and reliably.
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Lab products found in correlation
Tnf α, by Nanjing Jiancheng (3 mentions)
Il 6 elisa kit, by Nanjing Jiancheng (1 mentions)
Elx800, by Agilent Technologies (1 mentions)
Potassium oxonate, by Merck Group (1 mentions)
Allopurinol, by Merck Group (1 mentions)
Xanthine, by Merck Group (1 mentions)
Colchicine, by Merck Group (1 mentions)
Sod kit, by Nanjing Jiancheng (1 mentions)
Adenine, by Merck Group (1 mentions)
Synergy h1 microplate reader, by Agilent Technologies (1 mentions)
Jc 1 probe solution, by Beyotime (1 mentions)
Mitochondrial permeability transition pore assay kit, by Beyotime (1 mentions)
Enhanced mitochondrial membrane potential assay kit with jc 1, by Beyotime (1 mentions)
Il 1α, by Nanjing Jiancheng (1 mentions)
8 protocols using il 1β elisa kit
1
Subcutaneous Scaffold Implantation in Rats
2
Quantification of Inflammatory Cytokines
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The inflammatory cytokine levels of TNF-α, IL-6 and IL-10 in the blood plasma and intestinal tissues were quantified using ELISA kits (32 (link)). The following kits were used according to the manufacturer's protocol: TNF-α ELISA kit (cat. no. H052; Nanjing Jiancheng Bioengineering Institute), IL-1β ELISA kit (cat. no. H002; Nanjing Jiancheng Bioengineering Institute) and IL-6 ELISA kit (cat. no. H007; Nanjing Jiancheng Bioengineering Institute). Cytokine levels are expressed as pg/ml.
3
Measuring Cytokine Levels in Alveolar Lavage Fluid
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After collecting the alveolar lavage fluid, ELISA were performed according to the instructions for IL‐17A, tumour necrosis factor‐α (TNF‐α) and IL‐1β ELISA kits (H052 and H002, Nanjing Jiancheng Bioengineering institute). The antigen was solubilized in 50 mmol/L carbonate coating buffer (pH 9.6) to get an antigen concentration of 10‐20 µg/mL, and each well of the 96‐well microtitre plate was coated with 100 µL antigen solution at 4°C overnight. The coating buffer was discarded and each well was blocked for 1 hour with 50 µL 1% BSA at 37°C, followed by incubation with 100 µL of lavage solution with different dilutions at 37°C for 2 hours. Next, each well was incubated for 1 hour with 100 µL diluted HRP‐labelled secondary antibody at 37°C and then developed for 20 minutes. The absorbance at A405 was read using a microplate reader (BioTek ELx800, BioTek).
4
Quantification of Mucosal Immune Factors
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The levels of secretory IgA (sIgA ), IgM, IgG, tumor necrosis factor-α (TNF-α ), interferon-γ (IFN-γ ), and interleukin-1β (IL-1β ) in the supernatant obtained from intestinal mucosal homogenates were quantified with chicken-specific sIgA, IgM, IgG, TNF-α, IFN-γ, and IL-1β ELISA kits (Nanjing Jiancheng Bioengineering Institute), respectively. The measurement procedures were strictly in accordance with the protocols of manufacturer.
5
Kidney Injury Biomarker Assessment
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Tianjiang Pharmacology Co. Ltd (Jiangyin, China) provided Radix Aconiti Lateralis Praeparata, Concha Ostreae, Radix et Rhizoma Rhei, Herba Taraxaci, Flos Sophorae Immaturus, and Serissa serissoides Druce, whereas the China Institute for the Control of Pharmaceutical and Biological Products (Beijing, China) supplied all other related analytical reference (98% purity). Kidney injury factor-1 (KIM-1), serum creatinine (SCr), blood urea nitrogen (BUN), β2-microglobulin (β2-MG), malondialdehyde (MDA), superoxide dismutase (SOD), tumor necrosis factor alpha (TNF-α), and interleukin-1 β (IL-1β) ELISA kits were bought from the Nanjing JianCheng Bioengineering Institute (Nanjing, China). Abcam corporation (Cambridge, USA) supplied antibodies for aryl hydrocarbon receptor (AhR), anti-phospho-c-Jun-N-terminal kinase (JNK) cell signaling, nuclear factor-κB (NF-κB), antitropomyosin receptor kinase B (TrkB), and anti-brain-derived neurotrophic factor (BDNF). All other reagents and substances were kindly provided by Sigma Chem. Comp., (Sigma, Milan, Italy) unless otherwise stated.
6
Urate-Induced Renal Injury Model
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Intact plants of SM were purchased from Yi Chang in Hubei province, China. The samples were identified by authors (Prof. Ke-li Chen). Specimens of these plants were deposited in the herbarium, Hubei University of Chinese Medicine, China.
Sodium urate, potassium oxonate, adenine, allopurinol, colchicine, XOD, and xanthine were purchased from Sigma-Aldrich, USA. The UA, BUN, Cr, MPO, MDA, SOD kits, TNF-α, and IL-1β ELISA kits were purchased from Nanjing Jiancheng Bioengineering Institute, Nanjing City, China.
Sodium urate, potassium oxonate, adenine, allopurinol, colchicine, XOD, and xanthine were purchased from Sigma-Aldrich, USA. The UA, BUN, Cr, MPO, MDA, SOD kits, TNF-α, and IL-1β ELISA kits were purchased from Nanjing Jiancheng Bioengineering Institute, Nanjing City, China.
7
Murine Serum Cytokine and ROS Assay
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After induction model, mice were anaesthetized with Isoflurane (Induction: 3-5%, Maintenance: 1.5-3%), and peripheral blood were collected form submaxillary vein. Blood was centrifugated at1000 g at 4° C for 10 min and serum was collected and saved at -80° C for other experiment. Blood or cell samples were collected and used to measure ROS (E004-1-1), TNF-α (H052-1), IL-6 (H007-1-1), IL-18 (H015) and IL-1β (H002) levels using ROS, TNF-α, IL-6, IL-18 and IL-1β ELISA kits (Nanjing Jiancheng Biological Engineering Institute, Nanjing, China) following the manufacturer’s instructions.
1x 105/well cells were seeded into a 96-well plate and 100 μL JC-1 probe solution (C2006, Beyotime Biotechnology) was added into every well as previously described [34 (link)]. Absorbance was measured using a fluorescent reader (Synergy H1 Microplate Reader, Bio Tek, VT, USA).
1x 105/well cells were seeded into a 96-well plate and 100 μL JC-1 probe solution (C2006, Beyotime Biotechnology) was added into every well as previously described [34 (link)]. Absorbance was measured using a fluorescent reader (Synergy H1 Microplate Reader, Bio Tek, VT, USA).
8
Cytokine and Mitochondrial Assays
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Blood, tissue, or cell samples were collected and used to measure IL-1α, INF-γ, TNF-α, IL-6, and IL-1β levels using IL-1α, INF-γ, TNF-α, IL-6, and IL-1β ELISA kits (Nanjing Jiancheng Biological Engineering Institute, Nanjing, China) following the manufacturer's instructions. ROS production was evaluated by measuring ROS levels kits (S0033S, Beyotime Biotechnology) following the manufacturer's instructions. Calcein AM/CoCl2 assay and JC-1 disaggregation were evaluated by mitochondrial permeability transition pore assay kit (C2009S, Beyotime Biotechnology) and enhanced mitochondrial membrane potential assay kit with JC-1 (C2003S, Beyotime Biotechnology).
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