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TNF alpha ELISA Kit

TNF alpha Reactivity: Mouse Colorimetric Sandwich ELISA 60-6000 pg/mL Cell Culture Supernatant, Plasma, Serum
Catalog No. ABIN1979720
  • Target See all TNF alpha ELISA Kits
    TNF alpha (Tumor Necrosis Factor alpha (TNF alpha))
    Reactivity
    • 14
    • 12
    • 9
    • 5
    • 4
    • 4
    • 3
    • 2
    • 2
    • 2
    • 2
    • 2
    • 2
    • 1
    • 1
    • 1
    • 1
    • 1
    • 1
    Mouse
    Detection Method
    Colorimetric
    Method Type
    Sandwich ELISA
    Detection Range
    60-6000 pg/mL
    Minimum Detection Limit
    60 pg/mL
    Application
    ELISA
    Purpose
    Mouse TNF alpha ELISA Kit for cell culture supernatants, plasma, and serum samples.
    Sample Type
    Plasma, Cell Culture Supernatant, Serum
    Analytical Method
    Quantitative
    Specificity
    This ELISA kit shows no cross-reactivity with any of the cytokines tested: Mouse CD30, L CD30, T CD40, CRG-2, CTACK, CXCL16, Eotaxin , Eotaxin-2, Fas Ligand, Fractalkine, GCSF, GM-CFS, IFN- gamma, IGFBP-3, IGFBP-5, IGFBP-6, IL-1 alpha, IL-1 beta, IL-2, IL-3, IL-3 Rb, IL-4, IL-5, IL-6, IL-9, IL-10, IL-12 p40/p70, IL-12 p70, IL-13, IL-17, KC, Leptin R, LEPTIN(OB), LIX, L-Selectin, Lymphotactin, MCP-1, MCP-5, M-CSF, MIG, MIP-1 alpha, MIP-1 gamma, MIP-2, MIP-3 beta, MIP-3 alpha, PF-4, P-Selectin, RANTES, SCF, SDF-1 alpha, TARC, TCA-3, TECK, TIMP-1, TNF RI, TNF RII, TPO, VCAM-1, VEGF.
    Sensitivity
    < 60 pg/mL
    Characteristics
    • Strip plates and additional reagents allow for use in multiple experiments
    • Quantitative protein detection
    • Establishes normal range
    • The best products for confirmation of antibody array data
    Components
    • Pre-Coated 96-well Strip Microplate
    • Wash Buffer
    • Stop Solution
    • Assay Diluent(s)
    • Lyophilized Standard
    • Biotinylated Detection Antibody
    • Streptavidin-Conjugated HRP
    • TMB One-Step Substrate
    Material not included
    • Distilled or deionized water
    • Precision pipettes to deliver 2 μL to 1 μL volumes
    • Adjustable 1-25 μL pipettes for reagent preparation
    • 100 μL and 1 liter graduated cylinders
    • Tubes to prepare standard and sample dilutions
    • Absorbent paper
    • Microplate reader capable of measuring absorbance at 450nm
    • Log-log graph paper or computer and software for ELISA data analysis
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  • Application Notes
    Recommended Dilution for serum and plasma samples2 fold
    Sample Volume
    100 μL
    Plate
    Pre-coated
    Protocol
    1. Prepare all reagents, samples and standards as instructed in the manual.
    2. Add 100 μL of standard or sample to each well.
    3. Incubate 2.5 h at RT or O/N at 4 °C.
    4. Add 100 μL of prepared biotin antibody to each well.
    5. Incubate 1 h at RT.
    6. Add 100 μL of prepared Streptavidin solution to each well.
    7. Incubate 45 min at RT.
    8. Add 100 μL of TMB One-Step Substrate Reagent to each well.
    9. Incubate 30 min at RT.
    10. Add 50 μL of Stop Solution to each well.
    11. Read at 450 nm immediately.
    Reagent Preparation
    1. Bring all reagents and samples to room temperature (18 - 25 °C) before use.
      2. Sample dilution: If your samples need to be diluted, Assay Diluent A (Item D) should be used for dilution of serum/plasma samples. 1x Assay Diluent B (Item E) should be used for dilution of culture supernatants. Suggested dilution for normal serum/plasma: 2 fold. Please note that levels of the target protein may vary between different specimens. Optimal dilution factors for each sample must be determined by the investigator.
      3. Assay Diluent B should be diluted 5-fold with deionized or distilled water.
      4. Preparation of standard: Briefly spin the vial of Item C. Add 400 µL Assay Diluent A (for serum/plasma samples) or 1x Assay Diluent B (for cell culture medium and urine) into Item C vial to prepare a 50 ng/mL standard. Dissolve the powder thoroughly by a gentle mix. Add 120 µL TNF-alpha standard from the vial of Item C, into a tube with 880 µL Assay Diluent A or 1x Assay Diluent B to prepare a 6,000 pg/mL stock standard solution. Pipette 300 µL Assay Diluent A or 1x Assay Diluent B into each tube. Use the stock standard solution to produce a dilution series . Mix each tube thoroughly before the next transfer. Assay Diluent A or 1x Assay Diluent B serves as the zero standard (0 pg/mL). 300 µL 300 µL 300 µL 300 µL 300 µL 120 µL standard + 880 µL 300myl 6,000 3,000 1,500 750 375 187.5 93.75 0 pg/mL pg/mL pg/mL pg/mL pg/mL pg/mL pg/mL pg/mL
      5. If the Wash Concentrate (20x) (Item B) contains visible crystals, warm to room temperature and mix gently until dissolved. Dilute 20 ml of Wash Buffer Concentrate into deionized or distilled water to yield 400 ml of 1x Wash Buffer.
      6. Briefly spin the Detection Antibody vial (Item F) before use. Add 100 µL of 1x Assay Diluent B into the vial to prepare a detection antibody concentrate. Pipette up and down to mix gently (the concentrate can be stored at 4 °C for 5 days). The detection antibody concentrate should be diluted 80-fold with 1x Assay Diluent B and used in step 4 of Part VI Assay Procedure.
      7. Briefly spin the HRP-Streptavidin concentrate vial (Item G) and pipette up and down to mix gently before use. HRP-Streptavidin concentrate should be diluted 300-fold with 1x Assay Diluent B. For example: Briefly spin the vial (Item G) and pipette up and down to mix gently . Add 40 µL of HRP-Streptavidin concentrate into a tube with 12 ml 1x Assay Diluent B to prepare a 300-fold diluted HRP-Streptavidin solution (don't store the diluted solution for next day use). Mix well.
    Assay Procedure
    1. Bring all reagents and samples to room temperature (18 - 25 °C) before use. It is recommended that all standards and samples be run at least in duplicate.
      2. Add 100 µL of each standard (see Reagent Preparation step 2) and sample into appropriate wells. Cover well and incubate for 2.5 hours at room temperature or over night at 4 °C with gentle shaking.
      3. Discard the solution and wash 4 times with 1x Wash Solution. Wash by filling each well with Wash Buffer (300 myl) using a multi-channel Pipette or autowasher. Complete removal of liquid at each step is essential to good performance. After the last wash, remove any remaining Wash Buffer by aspirating or decanting. Invert the plate and blot it against clean paper towels.
      4. Add 100 µL of 1x prepared biotinylated antibody (Reagent Preparation step 6) to each well. Incubate for 1 hour at room temperature with gentle shaking.
      5. Discard the solution. Repeat the wash as in step
      6. Add 100 µL of prepared Streptavidin solution (see Reagent Preparation step 7) to each well. Incubate for 45 minutes at room temperature with gentle shaking.
      7. Discard the solution. Repeat the wash as in step
      8. Add 100 µL of TMB One-Step Substrate Reagent (Item H) to each well. Incubate for 30 minutes at room temperature in the dark with gentle shaking.
      9. Add 50 µL of Stop Solution (Item I) to each well. Read at 450 nm immediately.
    Calculation of Results

    Calculate the mean absorbance for each set of duplicate standards, controls and samples, and subtract the average zero standard optical density. Plot the standard curve on log-log graph paper or using Sigma plot software, with standard concentration on the x-axis and absorbance on the y-axis. Draw the best-fit straight line through the standard points.
    Typical Data: These standard curves are for demonstration only. A standard curve must be run with each assay. Mouse TNF-alpha concentration (pg/mL) O D =4 50 n m 0.01 0.1 1 10 Assay Buffer A 10 100 1,000 10,000 Mouse TNF-alpha concentration (pg/mL) O D =4 50 n m 0.01 0.1 1 10 Assay Buffer B 10 100 1,000 10,000
    Sensitivity: The minimum detectable dose of TNF-alpha is typically less than 60 pg/mL.
    Recovery: Recovery was determined by spiking various levels of Mouse TNF-alpha into Mouse serum, plasma and cell culture media. Mean recoveries are as follows: Sample Type Average % Recovery Range ( %) Serum 94.18 82-103 Plasma 93.82 83-102 Cell culture media 92.46 84-104
    Linearity: Sample Type Serum Plasma Cell culture media 1:2 Average % of Expected 93 94 92 Range ( %) 84-102 83-103 82-103 1:4 Average % of Expected 94 95 96 Range ( %) 82-103 84-104 83-104
    Reproducibility: Intra-Assay: CV<10 % Inter-Assay: CV<12 %

    Assay Precision
    Intra-Assay: CV< 10 % Inter-Assay: CV< 12 %
    Restrictions
    For Research Use only
  • Handling Advice
    Avoid repeated freeze-thaw cycles.
    Storage
    -20 °C
    Storage Comment
    The entire kit may be stored at -20°C for up to 1 year from the date of shipment. Avoid repeated freeze-thaw cycles. The kit may be stored at 4°C for up to 6 months. For extended storage, it is recommended to store at -80°C.
    Expiry Date
    6 months
  • Rasheed, Ahmed, Abdallah, El-Sayeh: "Nephro-toxic effects of intraperitoneally injected EGCG in diabetic mice: involvement of oxidative stress, inflammation and apoptosis." in: Scientific reports, Vol. 7, pp. 40617, (2018) (PubMed).

    Wang, Sun, Gou, Adams, Cui, Morgan, Strange, Wang: "α-1 Antitrypsin Enhances Islet Engraftment by Suppression of Instant Blood-Mediated Inflammatory Reaction." in: Diabetes, Vol. 66, Issue 4, pp. 970-980, (2017) (PubMed).

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    Singh, Kakalij, Kshirsagar, Kumar, Komakula, Diwan: "Cognitive effects of vanillic acid against streptozotocin-induced neurodegeneration in mice." in: Pharmaceutical biology, Vol. 53, Issue 5, pp. 630-6, (2015) (PubMed).

    Štofilová, Szabadosová, Hr?ková, Salaj, Bertková, Hijová, Strojný, Bomba: "Co-administration of a probiotic strain Lactobacillus plantarum LS/07 CCM7766 with prebiotic inulin alleviates the intestinal inflammation in rats exposed to N,N-dimethylhydrazine." in: International immunopharmacology, Vol. 24, Issue 2, pp. 361-8, (2015) (PubMed).

    Dey, Majhi, Mahanti, Dey, Bishayi: "In Vitro Anti-inflammatory and Immunomodulatory Effects of Ciprofloxacin or Azithromycin in Staphylococcus aureus-Stimulated Murine Macrophages are Beneficial in the Presence of Cytochalasin D." in: Inflammation, Vol. 38, Issue 3, pp. 1050-69, (2015) (PubMed).

    Ahmad, Attia, Bakheet, Zoheir, Ansari, Korashy, Abdel-Hamied, Ashour, Abd-Allah et al.: "Naringin attenuates the development of carrageenan-induced acute lung inflammation through inhibition of NF-?b, STAT3 and pro-inflammatory mediators and enhancement of I?B? and anti-inflammatory ..." in: Inflammation, Vol. 38, Issue 2, pp. 846-57, (2015) (PubMed).

    Bishayi, Bandyopadhyay, Majhi, Adhikary: "Effect of exogenous MCP-1 on TLR-2 neutralized murine macrophages and possible mechanisms of CCR-2/TLR-2 and MCP-1 signalling during Staphylococcus aureus infection." in: Immunobiology, Vol. 220, Issue 3, pp. 350-62, (2015) (PubMed).

    Ghosh, Bishayi: "Characterization of Toll-like receptor-4 (TLR-4) in the spleen and thymus of Swiss albino mice and its modulation in experimental endotoxemia." in: Journal of immunology research, Vol. 2015, pp. 137981, (2015) (PubMed).

    Yu, Yao, He, Tian: "Protection of MPTP-induced neuroinflammation and neurodegeneration by rotigotine-loaded microspheres." in: Life sciences, Vol. 124, pp. 136-43, (2015) (PubMed).

    Ghosh, Prakash, Manna, Bishayi: "Presence of toll like receptor-2 in spleen, lymph node and thymus of Swiss albino mice and its modulation by Staphylococcus aureus and bacterial lipopolysaccharide. ." in: Indian journal of experimental biology, Vol. 53, Issue 2, pp. 82-92, (2015) (PubMed).

    Kumar, Sahu, Kumar, Subuddhi, Maji, Jana, Gupta, Raffetseder, Lerm, Ghosh, van Loo, Beyaert, Gupta, Kundu, Basu: "MicroRNA let-7 modulates the immune response to Mycobacterium tuberculosis infection via control of A20, an inhibitor of the NF-?B pathway." in: Cell host & microbe, Vol. 17, Issue 3, pp. 345-56, (2015) (PubMed).

    Dey, Chaudhuri: "Immunomodulatory activity of Nerium indicum through inhibition of nitric oxide and cyclooxygenase activity and modulation of TH1/T H2 cytokine balance in murine splenic lymphocytes." in: Cytotechnology, (2015) (PubMed).

    Ghosh, Bishayi et al.: "Toll-like receptor 2 and 6 interdependency in the erosive stage of Staphylococcus aureus induced septic arthritis mediated by IFN-? and IL-6--A possible involvement of IL-17 in the progression of the ..." in: Immunobiology, Vol. 220, Issue 7, pp. 910-23, (2015) (PubMed).

    Zhong, Xu, Wang, Wang, Guan, Li, He, Liu, Zhou, Li, Wang, Liao: "Identification of anti-inflammatory constituents from Kalimeris indica with UHPLC-ESI-Q-TOF-MS/MS and GC-MS." in: Journal of ethnopharmacology, Vol. 165, pp. 39-45, (2015) (PubMed).

    Singla, Singla, Abdelli, Glass: "Fibroblast growth factor-9 enhances M2 macrophage differentiation and attenuates adverse cardiac remodeling in the infarcted diabetic heart." in: PLoS ONE, Vol. 10, Issue 3, pp. e0120739, (2015) (PubMed).

  • Target See all TNF alpha ELISA Kits
    TNF alpha (Tumor Necrosis Factor alpha (TNF alpha))
    Alternative Name
    TNF-alpha (TNF alpha Products)
    Synonyms
    DIF ELISA Kit, TNF-alpha ELISA Kit, TNFA ELISA Kit, TNFSF2 ELISA Kit, RATTNF ELISA Kit, Tnfa ELISA Kit, tnf ELISA Kit, TNF-a ELISA Kit, TNFalpha ELISA Kit, Tnfsf1a ELISA Kit, TNFa ELISA Kit, cTNF ELISA Kit, Tnf-alpha ELISA Kit, tnfa-like ELISA Kit, TNF-ALPHA ELISA Kit, dif ELISA Kit, tnfa ELISA Kit, xtnf ELISA Kit, tnfsf2 ELISA Kit, tnf-alpha ELISA Kit, Cachectin ELISA Kit, tumor necrosis factor ELISA Kit, tumor necrosis factor b (TNF superfamily, member 2) ELISA Kit, tumor necrosis factor alpha ELISA Kit, tumor necrosis factor a (TNF superfamily, member 2) ELISA Kit, TNF ELISA Kit, Tnf ELISA Kit, tnf ELISA Kit, tnfb ELISA Kit, tnf-alpha ELISA Kit, LOC103694380 ELISA Kit, tnfa ELISA Kit
    Background
    Tumor necrosis factor (Cachectin) (TNF-alpha) (Tumor necrosis factor ligand superfamily member 2) (TNF-a)
    Gene ID
    21926
    UniProt
    P06804
    Pathways
    NF-kappaB Signaling, Apoptosis, Caspase Cascade in Apoptosis, TLR Signaling, Cellular Response to Molecule of Bacterial Origin, Regulation of Leukocyte Mediated Immunity, Positive Regulation of Immune Effector Process, Production of Molecular Mediator of Immune Response, Positive Regulation of Endopeptidase Activity, Hepatitis C, Protein targeting to Nucleus, Inflammasome
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