The human umbilical vein endothelial cells (HUVEC, CRL-1730), human brain microvascular endothelial cells (HBMEC, CRL-3245), human aorta-vascular smooth muscle cells (HA-VSMC, CRL-1999), and human monocytes THP-1 (TIB-202) cell lines were purchased from the American Type Culture Collection (Manassas VA, USA); The oil red O solution was obtained from Beyotime Biotechnology (Shanghai, China); the shRNA lentivirus vector and the polybrene were obtained from the Cyagen Bioscience Inc., (Suzhou, China); IL-12, IL-6, and TNF-α ELISA kits were provided by Boster Biological Technology Co. Ltd. (Wuhan, China); Tissue Total cholesterol Assay Kit was from Applygen Technologies Inc. Beijing, China); PrimeScript RT reagent Kit, SYBR Premix DimerEraser™ (Perfect Real Time) assay kit, and the primers were purchased from Takara (Dalian, China); the primary antibodies p-TNK1 (D46E7), TNK1 (C44F9), p-STAT1 (58D6), STAT1 (D1K9Y), p-Tyk2 (Y1054), and Tyk2 (9312S) were purchased from CST (Danvers, MA, USA); the primary antibody for β-actin (AP0060) was purchased from Bioworld Technology (Nanjing, China); and the oxLDL was provided by the Peking Union-Biology, Co. Ltd (Beijing, China).
Tnf α elisa kit
Manufactured by Boster Bio
Sourced in China, United States
The TNF-α ELISA kit is a laboratory tool used to quantitatively measure the concentration of tumor necrosis factor-alpha (TNF-α) in various sample types. It employs the enzyme-linked immunosorbent assay (ELISA) technique to detect and quantify the target analyte.
Automatically generated - may contain errors
Lab products found in correlation
Human il 6 elisa kit, by Boster Bio (2 mentions)
Primescript rt reagent kit, by Takara Bio (1 mentions)
Ha vsmc, by American Type Culture Collection (1 mentions)
Hbmec, by American Type Culture Collection (1 mentions)
Thp 1 tib 202, by American Type Culture Collection (1 mentions)
Interleukin 6 (il 6), by Boster Bio (1 mentions)
Oil red o solution, by Beyotime (1 mentions)
Tissue total cholesterol assay kit, by Applygen (1 mentions)
Huvec crl 1730, by American Type Culture Collection (1 mentions)
Superoxide dismutase sod assay kit, by Nanjing Jiancheng (1 mentions)
Vit e, by Merck Group (1 mentions)
Lactate dehydrogenase ldh, by Nanjing Jiancheng (1 mentions)
Rpmi 1640, by Thermo Fisher Scientific (1 mentions)
Dextran sulfate sodium (dss), by Meilun (1 mentions)
Bicinchoninic acid bca protein assay kit, by Solarbio (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Occludin antibody, by Proteintech (1 mentions)
Fixable viability stain 780, by BD (1 mentions)
Gadph antibody, by Abcam (1 mentions)
Lipopolysaccharides (lps), by Merck Group (1 mentions)
19 protocols using tnf α elisa kit
1
Characterization of Endothelial and Smooth Muscle Cells
2
Antioxidant and Anti-inflammatory Effects
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Fish oil was purchased from Swiss company (Melbourne, Australia), corn oil from Shandong Sanxing corn technology Co., Ltd. (Shandong, China), and Vit E (α-tocopherol) from Sigma-Aldrich (St. Louis, USA). Fish oil, corn oil and Vit E were all certified as endotoxin free. IL-1β, IL-6, C-reactive protein (CRP), and TNF-α ELISA kits were purchased from Boster Biological Technology Co., Ltd. (Wuhan, China). 8-epi-prostaglandin F2α (8-epi-PGF2α) ELISA kit was purchased from Elabscience Biotechnology Co., Ltd. (Wuhan, China). Lactate dehydrogenase (LDH) and superoxide dismutase (SOD) assay kits were procured from Nanjing Jiancheng Bioengineering Institute (Nanjing, China).
3
Dextran Sulfate Sodium Colitis Model Protocol
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Dextran sodium sulfate
(DSS, molecular weight 36 000–50 000) was purchased
from Dalian Meilun Biotechnology Co., Ltd. (Dalian, China).l -Ergothioneine (EGT, ≥98%) was bought from Shanghai Aladdin
Biochemical Technology Co., Ltd. Fecal occult blood assay kit and
myeloperoxidase (MPO) assay kit were from Nanjing Jiancheng Bioengineering
Institute (Nanjing, China). Mouse IL-6, TNF-α ELISA kits, and
Goat anti-rabbit IgG were purchased from Boster Biological Technology
Co. Ltd. (Wuhan, China). The bicinchoninic acid (BCA) protein assay
kit was purchased from Solarbio Science & Technology Co., Ltd.
(Beijing, China). GADPH antibody was obtained from Abcam (Cambridge,
MA). The occludin antibody was purchased from Proteintech (Wuhan,
China). Fixable Viability Stain 780 from BD Pharmingen is useful for
discrimination of viable from nonviable cells. Antibodies for CD45,
CD3, CD4, CD11b, and CD16/CD32 (Fc Blocker) were bought from BD Pharmingen.
Antibody for F4/80 was from Biolegend. RPMI 1640 and fetal bovine
serum (FBS) were purchased from Gibco. Penicillin, streptomycin, and
glutamine were from Solarbio Life Technologies. Collagenase VIII and
DNAse I were from Sigma-Aldrich. Other chemicals were of analytical
grade.
(DSS, molecular weight 36 000–50 000) was purchased
from Dalian Meilun Biotechnology Co., Ltd. (Dalian, China).
Biochemical Technology Co., Ltd. Fecal occult blood assay kit and
myeloperoxidase (MPO) assay kit were from Nanjing Jiancheng Bioengineering
Institute (Nanjing, China). Mouse IL-6, TNF-α ELISA kits, and
Goat anti-rabbit IgG were purchased from Boster Biological Technology
Co. Ltd. (Wuhan, China). The bicinchoninic acid (BCA) protein assay
kit was purchased from Solarbio Science & Technology Co., Ltd.
(Beijing, China). GADPH antibody was obtained from Abcam (Cambridge,
MA). The occludin antibody was purchased from Proteintech (Wuhan,
China). Fixable Viability Stain 780 from BD Pharmingen is useful for
discrimination of viable from nonviable cells. Antibodies for CD45,
CD3, CD4, CD11b, and CD16/CD32 (Fc Blocker) were bought from BD Pharmingen.
Antibody for F4/80 was from Biolegend. RPMI 1640 and fetal bovine
serum (FBS) were purchased from Gibco. Penicillin, streptomycin, and
glutamine were from Solarbio Life Technologies. Collagenase VIII and
DNAse I were from Sigma-Aldrich. Other chemicals were of analytical
grade.
4
Cytokine Profiling in Brain Tissue
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Brain tissue in Fr1 area was homogenized and centrifuged at 12,000 rpm for 20 min. The supernatant was collected and stored at − 20 °C until use. IL-1β, IL-6 and TNF-αELISA kits (Boster Biotechnology Co., Ltd., Wuhan, China) were used for detection.
5
Cytokine and Neurotransmitter Regulation in Rats
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The rats were anaesthetized with pentobarbital, and the femoral artery was punctured to collect blood. The blood sample was placed in a heparin-coated tube and centrifuged (3500 rpm, 4 °C, 20 min). The plasma levels of IL-6, IL-1β, TNF-α, epinephrine, norepinephrine, dopamine and acetylcholine were measured 48 h after reperfusion. We used enzyme-linked immunosorbent assay (ELISA) kits to analyse the levels of each protein (#rat IL-6, #rat IL-1β and #rat TNF-α ELISA kit, Boster, China; # Rat Adrenaline, # Rat Norepinephrine, # Rat Dopamine and # Rat Acetylcholine, Langton, China) according to the manufacturer’s instructions (n = 6 rats/group).
6
Quantification of TNF-α and Intracellular ROS in RAW264.7 Cells
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TNF-α in the supernatant of the RAW264.7 cells treated by β
1-AA was quantified using the TNF-α ELISA kit (Boster, Pleasanton, USA) according to the manufacturer’s protocol. The intracellular ROS level in macrophages was quantified by using the 2,7-dichlorofluorescein diacetate (DCFH-DA; Sigma, St Louis, USA). The macrophages were seeded in 6-well plates at 1.6×10
5 cells/well and allowed to adhere overnight. Subsequently, the medium was replaced by fresh medium with different β
1-AA concentrations (10
–6 M, 10
–7 M, and 10
–8 M), negative IgG (10
–7 M), metoprolol (3× 10
–7 M), LPS (10 ng/mL; Sigma-Aldrich) or fresh medium alone and incubated for 24 h. The plates were washed twice with PBS and incubated with DCFH-DA (10 μM) at 37°C for 20 min. The mean fluorescence intensity (MFI) of the cells was determined at the best excitation wavelength of 485 nm and emission wavelength of 525 nm using the microplate reader.
1-AA was quantified using the TNF-α ELISA kit (Boster, Pleasanton, USA) according to the manufacturer’s protocol. The intracellular ROS level in macrophages was quantified by using the 2,7-dichlorofluorescein diacetate (DCFH-DA; Sigma, St Louis, USA). The macrophages were seeded in 6-well plates at 1.6×10
5 cells/well and allowed to adhere overnight. Subsequently, the medium was replaced by fresh medium with different β
1-AA concentrations (10
–6 M, 10
–7 M, and 10
–8 M), negative IgG (10
–7 M), metoprolol (3× 10
–7 M), LPS (10 ng/mL; Sigma-Aldrich) or fresh medium alone and incubated for 24 h. The plates were washed twice with PBS and incubated with DCFH-DA (10 μM) at 37°C for 20 min. The mean fluorescence intensity (MFI) of the cells was determined at the best excitation wavelength of 485 nm and emission wavelength of 525 nm using the microplate reader.
7
Cytokine Profiling in Murine Sera
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Concentrations of cytokines (TNF-α) were determined in serum aliquots from different groups of mice. TNF-α determination was carried out employing immunoassay and quimioluminiscence techniques using TNF-α ELISA kit (Boster biological technology Co. Ltd, CA, USA).
8
Quantification of IL-6 and TNF-α
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IL-6 and TNF-α concentration in the cultured supernatant were quantified by using the IL-6 ELISA kit, TNF-α ELISA Kit (Boster, Wuhan, China) according to the manufacturer’s instructions.
9
Evaluating Cytokine and Oxidative Stress
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The Human Interleukin 6 (IL-6) ELISA Kit (Invitrogen) and Human Tumor Necrosis Factor-α (TNF-α) ELISA Kit (Boster, Wuhan, China) were applied to evaluate the secretion levels of IL-6 and TNF-α in culture medium, respectively, as per the accompanying suggestion. The superoxide dismutase (SOD) activity and malondialdehyde (MDA) level in hPDLCs after the indicated transfection or/and LPS exposure were gauged by the SOD Activity Colorimetric Assay Kit (Abcam) and MDA Colorimetric Assay Kit (Elabsience, Wuhan, China), respectively, based on the manufacturers’ recommendations.
10
Evaluating Inflammatory Cytokines in Kidney Samples
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Whole kidneys or HK-2 cells were homogenized in a buffer containing 250 mM sucrose, 1 mM EDTA, 25 mM imidazole, 0.1 mM phenylmethylsulfonyl fluoride (PMSF) and 20 mM potassium phosphate buffer (pH 7.6), with 1% protease and phosphatase inhibitors (Roche Applied Science, IN, USA) at 4 °C. Tissue samples were centrifuged at 10,000 g for 30 min at 4 °C. The supernatant was used for determination of inflammatory cytokine concentrations and western blotting. IL-1β, TNF-α, MCP-1 and NOS-2 were determined by a mouse IL-1β ELISA Kit, a TNF-α ELISA Kit, an MCP-1 ELISA Kit (Boster, Wuhan, China) and an NOS-2 ELISA Kit (Elabscience, CA, USA). Procedures were performed according to the manufacturer’s instructions.
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