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|Antigen||Tumor Necrosis Factor (TNF) ELISA Kits|
|Reactivity||Mouse (Murine) Alternatives|
Kits with alternative reactivity to:
|Method Type||Sandwich ELISA|
|Detection Range||60-6000 pg/mL|
|Minimum Detection Limit||60 pg/mL|
|20 references available|
|Supplier||Log in to see|
Product Details Tumor Necrosis Factor ELISA KitTarget Details Application Details Handling References for Tumor Necrosis Factor Kit (ABIN625182) Images
|Purpose||Mouse TNF alpha ELISA Kit for cell culture supernatants, plasma, and serum samples.|
|Sample Type||Plasma, Cell Culture Supernatant, Serum|
|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|
|Material not included||
Target DetailsProduct Details Tumor Necrosis Factor ELISA Kit Application Details Handling References for Tumor Necrosis Factor Kit (ABIN625182) Images back to top
|Alternative Name||TNF-alpha (TNF ELISA Kit Abstract)|
|Background||Tumor necrosis factor (Cachectin) (TNF-alpha) (Tumor necrosis factor ligand superfamily member 2) (TNF-a)|
|Research Area||Virology, Atherosclerosis, Immunology, Receptors, Cytokines, Growth Factors, Innate Immunity, Cancer, Cardiovascular, Inflammation|
|Pathways||NF-kappaB Signaling, Apoptosis, Caspase Cascade in Apoptosis, 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|
Application DetailsProduct Details Tumor Necrosis Factor ELISA Kit Target Details Handling References for Tumor Necrosis Factor Kit (ABIN625182) Images back to top
|Application Notes||Recommended Dilution for serum and plasma samples2 fold|
|Sample Volume||100 μL|
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.
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|
HandlingProduct Details Tumor Necrosis Factor ELISA Kit Target Details Application Details References for Tumor Necrosis Factor Kit (ABIN625182) Images back to top
|Handling Advice||Avoid repeated freeze-thaw cycles.|
|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|
References for Tumor Necrosis Factor Kit (ABIN625182)Product Details Tumor Necrosis Factor ELISA Kit Target Details Application Details Handling Images back to top
|Product cited in:||
Yu, Yuan, Liu, Zhang, Wang, Zhang, Liu: "Delayed Administration of WP1066, an STAT3 Inhibitor, Ameliorates Radiation-Induced Lung Injury in Mice." in: Lung, Vol. 194, Issue 1, pp. 67-74, 2016
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