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CNTF ELISA Kit

CNTF Reaktivität: Human Colorimetric Sandwich ELISA 0.2-200 ng/mL Cell Culture Supernatant, Plasma, Serum
Produktnummer ABIN624959
  • Target Alle CNTF ELISA Kits anzeigen
    CNTF (Ciliary Neurotrophic Factor (CNTF))
    Reaktivität
    • 4
    • 3
    • 3
    • 2
    • 2
    • 2
    • 2
    • 1
    • 1
    • 1
    • 1
    • 1
    Human
    Nachweismethode
    Colorimetric
    Methodentyp
    Sandwich ELISA
    Detektionsbereich
    0.2-200 ng/mL
    Untere Nachweisgrenze
    0.2 ng/mL
    Applikation
    ELISA
    Verwendungszweck
    Human CNTF ELISA Kit for cell culture supernatants, plasma, and serum samples.
    Proben
    Plasma, Cell Culture Supernatant, Serum
    Analytische Methode
    Quantitative
    Spezifität
    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-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 beta, MIP-1 delta, PARC, PDGF, RANTES, SCF, TARC, TGF-beta, TIMP-1, TIMP-2, TNF-alpha, TNF-beta, TPO, VEGF.
    Sensitivität
    200 pg/mL
    Produktmerkmale
    • 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
    Bestandteile
    • 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
    Benötigtes Material
    • 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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  • Applikationshinweise
    Recommended Dilution for serum and plasma samples2 fold
    Probenmenge
    100 μL
    Plattentyp
    Pre-coated
    Protokoll
    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.
    Aufbereitung der Reagenzien
    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 supernantants and urine. Suggested dilution for normal serum/plasma: 2 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 200 ng/mL standard. Dissolve the powder thoroughly by a gentle mix. Pipette 300 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 ng/mL). Standard + 400 µL 150 µL 150 µL 150 µL 150 µL 150myl 200 66.67 22.22 7.407 2.469 0.823 0 ng/mL ng/mL ng/mL ng/mL ng/mL ng/mL ng/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 65-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 final 300 fold diluted HRP-Streptavidin solution (don't store the diluted solution for next day use). Mix well.
    Testdurchführung
    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.
    Ergebnisberechnung

    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. 0.1 1 10 100 1,000 Human CNTF concentration (ng/mL) O D =4 50 n m 0.01 0.1 1 10 Assay diluent A 0.1 1 10 100 1,000 Human CNTF concentration (ng/mL) O D =4 50 n m 0.01 0.1 1 10 Assay diluent B
    Sensitivity: The minimum detectable dose of CNTF is typically less than 0.2 ng/mL.
    Recovery: Recovery was determined by spiking various levels of human CNTF into human serum, plasma and cell culture media. Mean recoveries are as follows: Sample Type Average % Recovery Range ( %) Serum 92.35 82-102 Plasma 93.28 83-104 Cell culture media 95.45 84-104
    Linearity: Sample Type Serum Plasma Cell Culture Media 1:2 Average % of Expected 93 92 94 Range ( %) 84-103 83-105 84-105 1:4 Average % of Expected 94 95 92 Range ( %) 83-105 84-103 82-104
    Reproducibility: Intra-Assay: CV<10 % Inter-Assay: CV<12 %

    Testpräzision
    Intra-Assay: CV< 10 % Inter-Assay: CV< 12 %
    Beschränkungen
    Nur für Forschungszwecke einsetzbar
  • Handhabung
    Avoid repeated freeze-thaw cycles.
    Lagerung
    -20 °C
    Informationen zur Lagerung
    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.
    Haltbarkeit
    6 months
  • Sun, Mandai, Kamao, Hashiguchi, Shikamura, Kawamata, Sugita, Takahashi: "Protective Effects of Human iPS-Derived Retinal Pigmented Epithelial Cells in Comparison with Human Mesenchymal Stromal Cells and Human Neural Stem Cells on the Degenerating Retina in rd1 mice." in: Stem cells (Dayton, Ohio), Vol. 33, Issue 5, pp. 1543-53, (2015) (PubMed).

    Burnham, Louis, Magal, Varon: "Effects of ciliary neurotrophic factor on the survival and response to nerve growth factor of cultured rat sympathetic neurons." in: Developmental biology, Vol. 161, Issue 1, pp. 96-106, (1994) (PubMed).

    Davis, Yancopoulos: "The molecular biology of the CNTF receptor." in: Current opinion in cell biology, Vol. 5, Issue 2, pp. 281-5, (1993) (PubMed).

    Ip, Yancopoulos: "Ciliary neurotrophic factor and its receptor complex." in: Progress in growth factor research, Vol. 4, Issue 2, pp. 139-55, (1993) (PubMed).

    Arakawa, Sendtner, Thoenen: "Survival effect of ciliary neurotrophic factor (CNTF) on chick embryonic motoneurons in culture: comparison with other neurotrophic factors and cytokines." in: The Journal of neuroscience : the official journal of the Society for Neuroscience, Vol. 10, Issue 11, pp. 3507-15, (1990) (PubMed).

  • Target Alle CNTF ELISA Kits anzeigen
    CNTF (Ciliary Neurotrophic Factor (CNTF))
    Andere Bezeichnung
    CNTF (CNTF Produkte)
    Synonyme
    GPA ELISA Kit, CNTF ELISA Kit, HCNTF ELISA Kit, AI429687 ELISA Kit, ciliary neurotrophic factor ELISA Kit, CNTF ELISA Kit, Cntf ELISA Kit
    Hintergrund
    CNTF is found predominantly in peripheral nerve tissues. The main source appears to be myelin-associated Schwann cells in peripheral nerves and astrocytes in the central nervous system. In contrast to other Neurotrophins, CNTF appears to be expressed relatively late during ontogenesis. CNTF has been proposed to be a lesion factor that is released after nerve injuries and that, in combination with other factors, promotes the survival and the regeneration of neurons. The Human CNTF ELISA (Enzyme-Linked Immunosorbent Assay) kit is an in vitro enzyme-linked immunosorbent assay for the quantitative measurement of human CNTF in serum, plasma, cell culture supernatants and urine. This assay employs an antibody specific for human CNTF coated on a 96-well plate. Standards and samples are pipetted into the wells and CNTF present in a sample is bound to the wells by the immobilized antibody. The wells are washed and biotinylated anti-human CNTF 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 CNTF 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%.
    Gen-ID
    1270
    UniProt
    P26441
    Pathways
    JAK-STAT Signalweg
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