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Human TNF alpha SuperSet ELISA Kit

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SKU:
HUDC0136
$111 - $739

Description

Human TNF alpha SuperSet ELISA Kit

The Human TNF alpha SuperSet ELISA Kit is an ELISA kit for the detection of TNF-alpha in a range of sample types. SuperSet ELISA Kits are a range of specially designed ELISA Assays with the pharmaceutical and biotech research industries in mind. Focusing on the high quality monoclonal antibodies and purified standards, SuperSet ELISA Kits are key tools for immunology research. Each kit contains a highly optimised antibody pair as well as a purified recombinant protein for the detection of your analyte in cell culture supernatants. The SuperSet ELISA Kits can also be used for the detection of analytes in more complex matrices such as serum and plasma.

SuperSet ELISA Kit Key Features

  • Highly optimised monoclonal antibody pair for highly specific and sensitive analyte detection.
  • High quality purified recombinant protein to generate consistent standard curves.
  • Accuracy and reliability are guaranteed as all reagents have been validated according ISO 9001:2000 quality systems.
  • Recognises both Natural and Recombinant antigen Specificity.
  • No cross reactivity with other cytokines tested.
  • Standard Calibration to NIBSC.
  • ELISA Kits developed with pharmaceutical and biotech research sectors in mind.
Size: 1,5,10,15,20 x 96 Discovery
Alias: APC1 protein; Cachectin; Cachetin; DIF; TNF; TNF, monocyte-derived; tnfa; tnf-a; TNFalpha; TNF-alpha; TNF-alphacachectin; TNFATNF, macrophage-derived; TNFSF1A; TNFSF2; TNFSF2TNF superfamily, member 2; tumor necrosis factor (TNF superfamily, member 2); tumor necrosis factor alpha; Tumor necrosis factor ligand superfamily member 2; tumor necrosis factor; tumor necrosis factor-alpha
Assay Range: 25 pg/ml - 800 pg/ml
Sensitivity: 10 pg/ml
Target Species: Human
Specificity: Recognizes both natural and recombinant human TNF-a
Incubation: From sample to end 2h45
SampleType: Serum, Plasma, Cell culture supernatant
Sample Size: 100 µl
Cross Reactivity: No cross reactivity with other human cytokines. Cross reactivity with macac TNF-a. No interference with soluble TNFR I and TNFR II.
Kit Contents: SuperSet ELISA Kits include capture and biotinylated detection antibody, one standard per plate, Streptavidin-HRP, TMB, and detailed procedure including buffer composition.

Materials required but not provided

  • 96 well Microtiter plates: e.g. Nunc Maxisorp Cat # 468667 * Note: the use of ELISA plates which are not high affinity binding will result in lower performances.
  • Reconstitution Buffer: 1xPBS, 0.09% Azide *
  • Coating Buffer: 1xPBS, pH 7.2-7.4 *
  • Wash Buffer: 1xPBS, 0.05% Tween20 *
  • Blocking Buffer: 1xPBS, 5% BSA *
  • Standard Dilution Buffer: 1xPBS, 1% BSA *
  • Secondary Antibody Dilution Buffer: 1xPBS, 1% BSA * Note: Supplementation with 10% Animal Serum e.g. FCS for assaying serum, plasma or other body fluids samples may be necessary.
  • HRP Dilution Buffer: 1xPBS, 1% BSA, 0.1% Tween20 *
  • Stop Reagent: 1N Sulfuric Acid

* Available in an Accessory Pack for ELISA Set, ELISA Genie product RGDC00001, material for optimal performances and quantity provided for 5 plates.

  • Microtitre plate reader with appropriate filters 450nm required with optional 630nm reference filter
  • Microplate washer or wash bottle
  • 10, 50, 100, 200 and 1,000µl adjustable single channel micropipettes with disposable tips
  • 50-300µl multi-channel micropipette with disposable tips
  • Multichannel micropipette reagent reservoirs
  • Distilled water
  • Vortex mixer
  • Miscellaneous laboratory plastic and/or glass, if possible sterile

Technical Manual

ELISA Kit Technical ManualELISA Kit Technical Manual

Protocol

SuperSet ELISA Protocol

The below protocol is a sample protocol for a SuperSet ELISA using a biotinylated detection antibody and streptavidin-HRP. Sandwich ELISAs allow for the detection and quantification of an analyte in a sample by using known analyte concentrations as standards and plotting absorbance of known concentrations vs known standard concentrations. This allows the researcher to calculate the amount of analyte present in their sample. This can be very useful when looking for increases in protein concentration, phosphorylation of proteins or decreases on protein concentrations.

When diluting samples and reagents, they must be mixed completely and evenly. It is recommended to plot a standard curve for each test.

Sandwich Protocol

ELISA Kit Technical Manual

Procedure:

1. Add 100uL of diluted capture antibody to every well.
2. Cover with a plastic plate cover and incubate at 4°C overnight.
3. Remove the cover and wash the plate as follows: A) Aspirate the liquid from each well B) Dispense 0.3mL of washing solution into each well C) Aspirate the contents of each well D) Repeat step B and C
4. Add 100uL of blocking buffer to every well.
5. Cover with a plastic plate cover and incubate at room temperature (18-25°C) for 2 hours.
6. Wash plate X3.

Storage/Immediate Use:

7. For immediate use, please continue below. If you wish to store the coated and blocked plates for future use, bench dry each plate at room temperature (18-25°C) for 24 hours. Then store at 2-8°C in a sealed plastic bag with desiccant for up to 12 months.

8. Prepare the standard curve.
9. Add 100uL of each standard, sample & zero (standard dilution buffer) to appropriate wells in duplicate.
10. Add 50uL of diluted detection antibody into all wells.
11. Cover with a plastic plate cover and incubate at room temperature (18-25°C) for 4 hours.
12. Wash plate X2.
13. Add 100uL of Streptavidin HRP solution into each well.
14. Cover with a plastic plate cover and incubate at room temperature (18-25°C) for 30 minutes.
15. Wash plate X2.
16. Add 100uL ready-to-use TMB substrate solution into each well.
17. Incubate in the dark for 5-15 minutes at room temperature. Avoid direct exposure to light by wrapping the plate in aluminium foil.
18. Add 100uL Stop Reagent into each well.
19. Read at 450nm.
 
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Additional Information

Reactivity:
Human
ELISA Type:
SuperSet
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