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PDGF-AA Homodimer ELISA Kit

Reactivity: Rat Colorimetric Sandwich ELISA 90-6000 pg/mL Cell Culture Supernatant, Plasma, Serum
Catalog No. ABIN1980065
  • Target
    PDGF-AA Homodimer
    Reactivity
    • 3
    • 2
    • 2
    • 1
    Rat
    Detection Method
    Colorimetric
    Method Type
    Sandwich ELISA
    Detection Range
    90-6000 pg/mL
    Minimum Detection Limit
    90 pg/mL
    Application
    ELISA
    Purpose
    Rat PDGF-AA 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: rat CINC-2, CINC-3, CNTF, Fractalkine, IL-1 alpha, IL-1 beta, IL-4, IL-6, IL-10, GM-CSF, IFN-gamma, Leptin, Lix, MCP-1, MIP-3 alpha, beta- NGF, TIMP-1, TNF-alpha.
    Cross-Reactivity (Details)
    This ELISA kit shows no cross-reactivity with the following cytokines tested: rat CINC-2, CINC-3, CNTF, Fractalkine, IL-1alpha, IL-1beta, IL-4, IL-6, IL-10, GM-CSF, IFN-gamma, Leptin, Lix, MCP-1, MIP-3alpha, beta- NGF, TIMP-1, TNF-alpha.
    Sensitivity
    < 90 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
    Featured
    Discover our best selling PDGFA ELISA Kit
  • Application Notes
    Recommended Dilution for serum and plasma samples2 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. Suggested dilution for normal serum/plasma: 2 fold. RayBio®Rat PDGF-AA ELISA Kit Protocol 3 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) into Item C vial to prepare a 50 ng/mL standard. Dissolve the powder thoroughly by a gentle mix. Add 60 µL PDGF-AA 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 250 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). 250 µL 250 µL 250 µL 250 µL 250 µL 250myl 60 µL standard (50 ng/mL) + 440 µL 6,000 3,000 1,500 750.0 375.0 187.5 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. RayBio®Rat PDGF-AA ELISA Kit Protocol 4
      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 280-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 14 ml 1x Assay Diluent B to prepare a 280-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. RayBio®Rat PDGF-AA ELISA Kit Protocol 5
      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 Rat PDGF-AA concentration (pg/mL) O D =4 50 n m 0.01 0.1 1 10 10 100 1,000 10,000 Assay Diluent B Rat PDGF-AA concentration (pg/mL) O D =4 50 n m 0.01 0.1 1 10 10 100 1,000 10,000 RayBio®Rat PDGF-AA ELISA Kit Protocol 7
    Sensitivity: The minimum detectable dose of PDGF-AA is typically less than 90 pg/mL.
    Recovery: Recovery was determined by spiking various levels of rat PDGF-AA into serum, plasma and cell culture media. Mean recoveries are as follows: Sample Type Average % Recovery Range ( %) Serum 100.0 91-108 Plasma 101.6 84-114 Cell culture media 130.4 122-138
    Linearity: Sample Type Serum Plasma Cell Culture Media 1:2 Average % of Expected 107.7 101.9 106.9 Range ( %) 96-115 92-110 99-114 1:4 Average % of Expected 108.5 75.87 82.93 Range ( %) 99-116 67-83 73-91 RayBio®Rat PDGF-AA ELISA Kit Protocol 8
    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
    PDGF-AA Homodimer
    Alternative Name
    PDGF-AA
    Synonyms
    PDGF-A ELISA Kit, PDGF1 ELISA Kit, AA409063 ELISA Kit, CD142 ELISA Kit, Cf-3 ELISA Kit, Cf3 ELISA Kit, TF ELISA Kit, PDGFACP ELISA Kit, platelet derived growth factor subunit A ELISA Kit, coagulation factor III ELISA Kit, platelet derived growth factor, alpha ELISA Kit, PDGFA ELISA Kit, F3 ELISA Kit, Pdgfa ELISA Kit
    Background
    Pdgfa protein (Platelet derived growth factor, alpha, isoform CRA_b)
    Gene ID
    25266
    UniProt
    Q6P7C3
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