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TRAIL ELISA Kit

TNFSF10 Reactivity: Human Colorimetric Sandwich ELISA 32-2000 pg/mL Cell Lysate, Plasma, Serum
Catalog No. ABIN924801
  • Target See all TRAIL (TNFSF10) ELISA Kits
    TRAIL (TNFSF10) (Tumor Necrosis Factor (Ligand) Superfamily, Member 10 (TNFSF10))
    Reactivity
    • 8
    • 5
    • 3
    • 3
    • 2
    • 1
    • 1
    • 1
    • 1
    • 1
    • 1
    • 1
    Human
    Detection Method
    Colorimetric
    Method Type
    Sandwich ELISA
    Detection Range
    32-2000 pg/mL
    Minimum Detection Limit
    32 pg/mL
    Application
    ELISA
    Purpose
    The OmniKine? Human TRAIL/ApoII Ligand ELISA Kit contains the components necessary for quantitative determination of natural or recombinant Human TRAIL/ApoII Ligand concentrations within any experimental sample including cell lysates, serum and plasma. This particular immunoassay utilizes the quantitative technique of a "Sandwich" Enzyme-Linked Immunosorbent Assay (ELISA) where the target protein (antigen) is bound in a "sandwich" format by the primary capture antibodies coated to each well-bottom and the secondary detection antibodies added subsequently by the investigator. The capture antibodies coated to the bottom of each well are specific for a particular epitope on Human TRAIL/ApoII Ligand while the user-added detection antibodies bind to epitopes on the captured target protein. Amid each step of the procedure, a series of wash steps must be performed to ensure the elimination of non- specific binding between proteins to other proteins or to the solid phase. After incubation and "sandwiching" of the target antigen, a peroxidase enzyme is conjugated to the constant heavy chain of the secondary antibody (either covalently or via Avidin/Streptavidin-Biotin interactions), allowing for a colorimetric reaction to ensue upon substrate addition. When the substrate TMB (3, 3', 5, 5'-Tetramethylbenzidine) is added, the reaction catalyzed by peroxidase yields a blue color that is representative of the antigen concentration. Upon sufficient color development, the reaction can be terminated through addition of Stop Solution (2 N Sulfuric Acid) where the color of the solution will turn yellow. The absorbance of each well can then be read by a spectrophotometer, allowing for generation of a standard curve and subsequent determination of protein concentration.
    Brand
    OmniKine™
    Sample Type
    Cell Lysate, Serum, Plasma
    Analytical Method
    Quantitative
    Specificity
    The Human TRAIL/ApoII Ligand ELISA Kit allows for the detection and quantification of endogenous levels of natural and/or recombinant Human TRAIL/ApoII Ligand proteins.
    Cross-Reactivity (Details)
    The Human sTRAIL ELISA is capable of recognizing both recombinant and naturally produced Human sTRAIL proteins. The antigens listed below were tested at 50 ng/mL and exhibited less than 1% significant cross reactivity or interference. Human: TNFβ, sRANKR The antigens listed below were tested at 50 ng/mL and did not exhibit significant cross reactivity or interference. Human: 4-1BBL, 4-1BBR, BAFF, CD40L, FasL, OPG, sRANKL, TNFα, sTNF-RI, sTNF-RII, sTRAIL-RI, sTRAIL-RII, TWEAK, TWEAKR Murine: TNFα Rat: TNF
    Characteristics
    The Human TRAIL/ApoII Ligand ELISA Kit allows for the detection and quantification of endogenous levels of natural and/or recombinant Human TRAIL/ApoII Ligand proteins within the range of 32-2000 pg/mL.
    Components
    • Microstrips Coated w / Capture Antibody: 12 x 8-Well Microstrips
    • Protein Standard: Lyophilized (100 ng), Red container
    • Biotinylated Detection Antibody: Lyophilized, Yellow container
    • 400x Streptavidin-HRP: 30 μL, Blue container
    • Wash Buffer (10x): 50 mL, Clear containter
    • Assay Diluent: 50 mL, Clear container
    • Ready-to-Use Substrate: 12 mL, Brown container
    • Stop Solution: 12 mL, Clear container
    • Adhesive Plate Sealers: 4 Sheets
    • Technical Manual 1 Manual
    Material not included
    The following materials and/or equipment are NOT provided in this kit but are necessary to successfully conduct the experiment:
    Microplate reader able to measure absorbance at 450 nm (with correction wavelength set to 540 nm or 570 nm)
    Micropipettes with capability of measuring volumes ranging from 1 μl to 1 mL
    Deionized or sterile water
    Squirt bottle, manifold dispenser, multichannel pipette reservoir or automated microplate washer
    Graph paper or computer software capable of generating or displaying logarithmic functions
    Absorbent paper or vacuum aspirator
    Test tubes or microfuge tubes capable of storing ≥1 mL
    Bench
    top centrifuge (optional)
    Bench
    top vortex (optional)
    Orbital shaker (optional)
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  • Plate
    Pre-coated
    Protocol
    This particular immunoassay utilizes the quantitative technique of a Sandwich Enzyme-Linked Immunosorbent Assay (ELISA) where the target protein (antigen) is bound in a sandwich format by the primary capture antibodies coated to each well-bottom and the secondary detection antibodies added subsequently by the investigator. The capture antibodies coated to the bottom of each well are specific for a particular epitope on the Human sTRAIL cytokine while the user-added detection antibodies bind to epitopes on the captured target protein. Amid each step of the procedure, a series of wash steps must be performed to ensure the elimination of non-specific binding between proteins to other proteins or to the solid phase. After incubation and sandwiching of the target antigen, a peroxidase enzyme is conjugated to the constant heavy chain of the secondary antibody (either covalently or via Avidin/Streptavidin-Biotin interactions), allowing for a colorimetric reaction to ensue upon substrate addition. When the substrate TMB (3, 3’, 5, 5’- Tetramethylbenzidine) is added, the reaction catalyzed by peroxidase yields a blue color that is representative of the antigen concentration. Upon sufficient color development, the reaction can be terminated through addition of Stop Solution (2 N Sulfuric Acid) where the color of the solution will turn yellow. The absorbance of each well can then be read by a spectrophotometer, allowing for generation of a standard curve and subsequent determination of protein concentration.
    Sample Preparation

    If samples are to be used within 24 hours, aliquot and store at 4 °C. If samples are to be used over a long period of time, aliquot and store between -20 °C and -80 °C, depending on the duration of storage.
    Note: Samples containing a visible precipitate or pellet must be clarified prior to use in the assay.
    Caution: Avoid repeated freeze/thaw cycles to prevent loss of biological activity of proteins in experimental samples.

    • Cell Lysate and Supernatants:
      Remove large cell components via centrifugation and perform the assay. Cell lysates and supernatants require a dilution using Assay Diluent. A serial dilution may be performed to determine a suitable dilution factor for the sample. For future use of the sample, follow the sample storage guidelines stated above.
    • Serum:
      Allow samples to clot in a serum separator tube (SST) for 30 minutes. After sufficient clotting, centrifuge at 1000 x g for 15 minutes and remove serum from SST in preparation for the assay. Serum samples require at least a 1:50 dilution using Assay Diluent. For future use of the sample, follow the storage guidelines above.
    • Plasma:
      Use heparin, citrate or EDTA as an anticoagulant to gather plasma from original biological sample. After collection of the plasma, centrifuge for 15 minutes at 1000 x g. This step must be performed within 30 minutes of plasma collection. Plasma samples require at least a 1:50 dilution using Assay Diluent. Afterwards, perform the assay or for future use of the sample, follow the storage guidelines stated above.

    Assay Procedure

    Note: If possible, all incubation steps should be performed on an orbital shaker to equilibrate solutions when added to the microplate wells. Also, all provided solutions should be at ambient temperature prior to use.
    Note: Avoid adding solutions into wells at an angle, always keep pipette tip perpendicular to plate bottom.

    Reconstitution of Provided Materials:

      1. Reconstitute the Biotin-Conjugated Detection Antibody in 67 µL of ddH₂O for a concentration of 180 µg/ml.
      2. Reconstitute the Protein Standard in 100 µL of ddH₂O for a concentration of 340 ng/ml.
      3. Dilute the 50 mL of 10x Wash Buffer in 450 mL of ddH2O for 500 mL of 1x Wash Buffer.
    Addition of Known Standard and Unknown Sample to Immunoassay:
      The OmniKine™ Human CD163 ELISA Kit allows for the detection and quantification of endogenous levels of natural and/or recombinant Human CD163 proteins

    Calculation of Results

    Generation of Standard Curve and Interpretation of Data
    1. Average the duplicate or triplicate readings for each standard, control and sample and subtract the average zero standard optical density.
    2. Generate a standard curve by using Microsoft Excel or other computer software capable of establishing a 4- Parameter Logistic (4-PL) curve fit. If using Excel or an alternative graphing tool, plot the average optical density values in absorbance units (y-axis) against the known standard concentrations in pg/ml (x-axis). Note: Only use the values in which a noticeable gradient can be established. Afterwards, generate a best fit curve or trend-line through the plotted points via regression analysis.

    Restrictions
    For Research Use only
  • Precaution of Use
    Reagents provided in this kit may be harmful if ingested, inhaled or absorbed through the skin. Please carefully review the MSDS for each reagent before conducting the experiment.
    Stop Solution contains 2 N Sulfuric Acid (H2SO4) and is an extremely corrosive agent. Please wear proper eye, hand and face protection when handling this material. When the experiment is finished, be sure to rinse the plate with copious amounts of running water to dilute the Stop Solution prior to disposing the plate.
    Handling Advice
    This ELISA kit is intended for research purposes only, NOT diagnostic or clinical procedures of any kind.
    Materials included in this kit should NOT be used past the expiration date on the kit label.
    Reagents or substrates included in this kit should NOT be mixed or substituted with reagents or substrates from any other kits.
    Variations in pipetting technique, washing technique, operator laboratory technique, kit age, incubation time or temperature may cause differences in binding affinity of the materials provided.
    The assay is designed to eliminate interference and background by other cellular macromolecules or factors present within any biological samples. However, the possibility of background noise cannot be fully excluded until all factors have been tested using the assay kit.

    Reagents provided in this kit may be harmful if ingested, inhaled or absorbed through the skin. Please carefully review the MSDS for each reagent before conducting the experiment.
    Stop Solution contains 2 N Sulfuric Acid (H2SO4) and is an extremely corrosive agent. Please wear proper eye, hand and face protection when handling this material. When the experiment is finished, be sure to rinse the plate with copious amounts of running water to dilute the Stop Solution prior to disposing the plate.
    Storage
    4 °C
    Storage Comment
    Note: If used frequently, reagents may be stored at 4 °C.
    • Unopened Kits: Store at 4 °C for 6 months.
    • Microstrips Coated w/ Capture Antibody, 400x Streptavidin-HRP Wash Buffer (10x), Assay Diluent Ready-to-Use Substrate, Stop Solution: 6 Months at 4 °C
    • Protein Standard, Biotinylated Detection Antibody: Lyophilized: 6 Months (if Reconstituted: 1 Month) at 4 °C
  • Target See all TRAIL (TNFSF10) ELISA Kits
    TRAIL (TNFSF10) (Tumor Necrosis Factor (Ligand) Superfamily, Member 10 (TNFSF10))
    Abstract
    TNFSF10 Products
    Synonyms
    APO2L ELISA Kit, Apo-2L ELISA Kit, CD253 ELISA Kit, TL2 ELISA Kit, TRAIL ELISA Kit, A330042I21Rik ELISA Kit, AI448571 ELISA Kit, APO-2L ELISA Kit, Ly81 ELISA Kit, Trail ELISA Kit, tl2 ELISA Kit, apo2l ELISA Kit, cd253 ELISA Kit, trail ELISA Kit, apo-2l ELISA Kit, trail1 ELISA Kit, xtrail1 ELISA Kit, tnfsf10l2 ELISA Kit, zgc:92320 ELISA Kit, TNLG6A ELISA Kit, TNF superfamily member 10 ELISA Kit, tumor necrosis factor (ligand) superfamily, member 10 ELISA Kit, tumor necrosis factor superfamily member 10 ELISA Kit, TNF superfamily member 10 S homeolog ELISA Kit, TNFSF10 ELISA Kit, Tnfsf10 ELISA Kit, tnfsf10.S ELISA Kit, tnfsf10 ELISA Kit
    Background
    Human sTRAIL or Soluble TNF-Related Apoptosis-Inducing Ligand, also known as Tumor Necrosis Factor Ligand Superfamily Member 10, is a 281 amino acid cytokine protein encoded by the TNFSF10 gene located at locus 3q26 on chromosome 3. The sTRAIL cytokine is able to induce apoptosis through binding with TNFRSF10A/TRAILR1, TNFRSF10B/TRAILR2, TNFRSF10C/TRAILR3, TNFRSF10D/TRAILR4 and possibly also to TNFRSF11B/OPG. Furthermore, its activity may be modulated by binding to the decoy receptors TNFRSF10C/TRAILR3, TNFRSF10D/TRAILR4 and TNFRSF11B/OPG that cannot induce apoptosis. Essentially, sTRAIL preferentially induces apoptosis in transformed and tumor cells, but does not appear to kill normal cells although it is expressed at a significant level in most normal tissues. The binding of this protein to its receptors has been shown to trigger the activation of MAPK8/JNK, caspase-8, and caspase-3. Source: Entrez Gene: TNFSF10 tumor necrosis factor (ligand) superfamily, member 10 [Homo sapiens], Swiss-Prot: P50591
    Gene ID
    8743
    UniProt
    P50591
    Pathways
    Apoptosis, Positive Regulation of Endopeptidase Activity
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