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Adipsin ELISA Kit

CFD Reactivity: Human Colorimetric Sandwich ELISA 4-6000 pg/mL Cell Culture Supernatant, Plasma, Serum
Catalog No. ABIN1979914
  • Target See all Adipsin (CFD) ELISA Kits
    Adipsin (CFD) (Complement Factor D (CFD))
    Reactivity
    • 4
    • 2
    • 2
    • 2
    • 1
    • 1
    Human
    Detection Method
    Colorimetric
    Method Type
    Sandwich ELISA
    Detection Range
    4-6000 pg/mL
    Minimum Detection Limit
    4 pg/mL
    Application
    ELISA
    Purpose
    Human Adipsin (Complement Factor D) 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 the following 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-6, IL-7, IL-8, IL-9, IL-11, IL-12 p70, IL-12 p40, IL-13, IL-15, I-309, IP-10, G-CSF, GM-CSF, IFN-gamma, Leptin (OB), 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
    < 4 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 samples10,000 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 cell culture supernates/urine. Suggested dilution for normal serum/plasma: 10,000 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 before use.
      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 supernates/urine) into Item C vial to prepare a 50 ng/mL standard. Dissolve the powder thoroughly by a gentle mix. Add 60 µL Adipsin standard (50 ng/mL) from the vial of Item C, into a tube with 440 µL Assay Diluent A or 1x Assay Diluent B to prepare a 6,000 pg/mL standard solution. Pipette 400 µL Assay Diluent A or 1x Assay Diluent B into each tube. Use the 6,000 pg/mL 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). 60 µL standard + 440 µL 200 µL 200 µL 200 µL 200 µL 200 µL 200myl 6,000 2,000 666.7 222.2 74.07 24.69 8.23 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 50 µL of HRP-Streptavidin concentrate into a tube with 15 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. Assay Diluent A Human Adipsin concentration (pg/mL) 1 10 100 1000 10000 O D =4 50 n m 0.01 0.1 1 10 Assay Diluent B Human Adipsin 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 Adipsin is typically less than 4 pg/mL.
    Recovery: Recovery was determined by human Adipsin into human serum, plasma and cell culture media. Mean recoveries are as follows: Sample Type Average % Recovery Range ( %) Serum 130.3 118-139 Plasma 131.1 107-140 Cell culture media 126.4 115-133
    Linearity: Sample Type Serum Plasma Cell Culture Media 1:2 Average % of Expected 104.3 126.3 122.1 Range ( %) 93-114 107-135 111-130 1:4 Average % of Expected 95.46 130.7 126.5 Range ( %) 85-104 108-137 117-135
    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
  • Target See all Adipsin (CFD) ELISA Kits
    Adipsin (CFD) (Complement Factor D (CFD))
    Alternative Name
    Adipsin (CFD Products)
    Synonyms
    ADIPSIN ELISA Kit, ADN ELISA Kit, DF ELISA Kit, PFD ELISA Kit, Adn ELISA Kit, Df ELISA Kit, EVE ELISA Kit, CFD ELISA Kit, adipsin ELISA Kit, pfd ELISA Kit, cfdl ELISA Kit, wu:fb61f12 ELISA Kit, zgc:109940 ELISA Kit, 10.5 ELISA Kit, 10.9 ELISA Kit, 14.10 ELISA Kit, 20.35 ELISA Kit, CG2328 ELISA Kit, Dmel\\CG2328 ELISA Kit, E(eve) ELISA Kit, Eve ELISA Kit, F ELISA Kit, V ELISA Kit, VI ELISA Kit, eve2 ELISA Kit, even ELISA Kit, l(2)46CFg ELISA Kit, l(2)46CFh ELISA Kit, l(2)46CFj ELISA Kit, l(2)46CFp ELISA Kit, l(2)46Ce ELISA Kit, l(2)46Cg ELISA Kit, complement factor D ELISA Kit, complement factor D (adipsin) ELISA Kit, complement factor D (adipsin) L homeolog ELISA Kit, even skipped ELISA Kit, CFD ELISA Kit, Cfd ELISA Kit, cfd.L ELISA Kit, cfd ELISA Kit, eve ELISA Kit
    Background
    The Human Adipsin/Factor D ELISA (Enzyme-Linked Immunosorbent Assay) kit is an in vitro enzyme-linked immunosorbent assay for the quantitative measurement of human Adipsin in serum, plasma, cell culture supernatants and urine. This assay employs an antibody specific for human Adipsin coated on a 96-well plate. Standards and samples are pipetted into the wells and Adipsin present in a sample is bound to the wells by the immobilized antibody. The wells are washed and biotinylated anti-human Adipsin 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 Adipsin 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%.
    Gene ID
    1675
    UniProt
    P00746
    Pathways
    Complement System
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