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|Antigen||Tumor Necrosis Factor Receptor Superfamily, Member 1B (TNFRSF1B) ELISA Kits|
Kits with alternative reactivity to:
|Method Type||Sandwich ELISA|
|Detection Range||5-2000 pg/mL|
|Minimum Detection Limit||5 pg/mL|
|1 reference available|
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Product Details TNFRSF1B ELISA KitTarget details Application Details Handling ProductDetails: References for TNFRSF1B Kit (ABIN625090) Images
|Purpose||Human TNF RII (TNFRSF1B) 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: Human Angiogenin, BDNF, BLC, ENA-78, FGF-4, IL-1 alpha, IL-1 beta, IL-2, IL-3, IL-4, IL-5, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12 p70, IL-12 p40, IL-13, IL-15, I-309, IP-10, G-CSF, GM-CSF, IFN-gamma, Leptin, MCP-1, MCP-2, MCP-3, MDC, MIP-1 alpha, MIP-1 beta, MIP-1 delta, PARC, PDGF, RANTES, SCF, TARC, TGF-beta, TIMP-1, TIMP-2, TNF-alpha, TNF-beta, TPO, VEGF.|
|Sensitivity||< 5 pg/mL|
|Material not included||
Target detailsProduct Details TNFRSF1B ELISA Kit Application Details Handling ProductDetails: References for TNFRSF1B Kit (ABIN625090) Images back to top
|Alternative Name||TNF RII (TNFRSF1B ELISA Kit Abstract)|
|Background||The Human sTNF RII ELISA (Enzyme-Linked Immunosorbent Assay) kit is an in vitro enzyme-linked immunosorbent assay for the quantitative measurement of human sTNF RII in serum, plasma, cell culture supernatants and urine. This assay employs an antibody specific for human sTNF RII coated on a 96-well plate. Standards and samples are pipetted into the wells and sTNF RII present in a sample is bound to the wells by the immobilized antibody. The wells are washed and biotinylated anti-human sTNF RII antibody is added. After washing away unbound biotinylated antibody, HRP-conjugated streptavidin is pipetted to the wells. The wells are again washed, a TMB substrate solution is added to the wells and color develops in proportion to the amount of sTNF RII bound. The Stop Solution changes the color from blue to yellow, and the intensity of the color is measured at 450 nm. Reproducibility: Intra-Assay: CV<10% Inter-Assay: CV<12%.|
|Pathways||NF-kappaB Signaling, Apoptosis, Cellular Response to Molecule of Bacterial Origin, Hepatitis C|
Application DetailsProduct Details TNFRSF1B ELISA Kit Target details Handling ProductDetails: References for TNFRSF1B Kit (ABIN625090) Images back to top
|Application Notes||Recommended Dilution for serum and plasma samples5 fold - 10 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 and urine. Suggested dilution for normal serum/plasma: 5-10 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 and then 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 40 µL sTNF RII standard from the vial of Item C, into a tube with 960 µL Assay Diluent A or 1x Assay Diluent B to prepare a 2,000 pg/mL stock standard solution. Pipette 400 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). 200 µL 200myl 200 µL 200 µL 200 µL 200 µL 40 µL standard + 960 µL 2000 666.7 222.2 74.07 24.69 8.23 2.74 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 500-fold with 1x Assay Diluent B. For example: Briefly spin the vial (Item G) and pipette up and down to mix gently . Add 20 µL of HRP-Streptavidin concentrate into a tube with 10 ml 1x Assay Diluent B to prepare a 500-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. Assay Diluent A Human sTNFRII concentration (pg/mL) 1 10 100 1000 10000 O D =4 50 n m 0.01 0.1 1 10 Assay Diluent B Human sTNFRII concentration (pg/mL) 1 10 100 1000 10000 O D =4 50 n m 0.01 0.1 1 10
Sensitivity: The minimum detectable dose of sTNF RII is typically less than 5 pg/mL.
Recovery: Recovery was determined by spiking various levels of human sTNF RII into human serum, plasma and cell culture media. Mean recoveries are as follows: Sample Type Average % Recovery Range ( %) Serum 104.92 95-115 Plasma 105.49 94-118 Cell culture media 102.65 91-112
Linearity: Sample Type Serum Plasma Cell Culture Media 1:2 Average % of Expected 102 104 101 Range ( %) 88-111 89-113 86-108 1:4 Average % of Expected 99 102 97 Range ( %) 85-106 86-108 84-107
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 TNFRSF1B ELISA Kit Target details Application Details ProductDetails: References for TNFRSF1B Kit (ABIN625090) 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|
ProductDetails: References for TNFRSF1B Kit (ABIN625090)Product Details TNFRSF1B ELISA Kit Target details Application Details Handling Images back to top
|Product cited in:||
Cuchacovich, Hagan, Khan, Richert, Espinoza: "Tumor necrosis factor-alpha (TNF-α)-blockade-induced hepatic sarcoidosis in psoriatic arthritis (PsA): case report and review of the literature." in: Clinical rheumatology, Vol. 30, Issue 1, pp. 133-7, 2011
ImagesProduct Details TNFRSF1B ELISA Kit Target details Application Details Handling ProductDetails: References for TNFRSF1B Kit (ABIN625090) back to top