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SERPINA1 ELISA Kit

SERPINA1 Reaktivität: Human Colorimetric Sandwich ELISA 0.391-100 ng/mL Cell Culture Cells, Cerebrospinal Fluid, Milk, Saliva, Urine
Produktnummer ABIN612659
  • Target Alle SERPINA1 ELISA Kits anzeigen
    SERPINA1 (serpin Peptidase Inhibitor, Clade A (Alpha-1 Antiproteinase, Antitrypsin), Member 1 (SERPINA1))
    Reaktivität
    • 5
    • 4
    • 3
    • 2
    • 2
    • 2
    • 1
    • 1
    • 1
    • 1
    • 1
    • 1
    • 1
    Human
    Nachweismethode
    Colorimetric
    Methodentyp
    Sandwich ELISA
    Detektionsbereich
    0.391-100 ng/mL
    Untere Nachweisgrenze
    0.391 ng/mL
    Applikation
    ELISA
    Verwendungszweck
    The AssayMax™ Human alpha-1-Antitrypsin ELISA (Enzyme-Linked Immunosorbent Assay) Kit is designed for detection of A1AT in human plasma, serum, urine, saliva, milk, CSF, and cell culture samples. This assay employs a quantitative sandwich enzyme immunoassay technique that measures human A1AT in approximately 4 hours. A polyclonal antibody specific for human A1AT has been pre-coated onto a 96-well microplate with removable strips. A1AT in standards and samples is sandwiched by the immobilized antibody and a biotinylated polyclonal antibody specific for human A1AT, which is recognized by a streptavidin-peroxidase (SP) conjugate. All unbound material is washed away and a peroxidase enzyme substrate is added. The color development is stopped and the intensity of the color is measured.
    Marke
    AssayMax™
    Proben
    Cell Culture Cells, Cerebrospinal Fluid, Milk, Saliva, Urine
    Analytische Methode
    Quantitative
    Spezifität
    If cell culture supernatants contains 10% FBS, the minimum detectable dose of human A1AT will be 2 ng/ml.
    Bestandteile
    Human alpha-1-Antitrypsin Microplate: A 96-well polystyrene microplate (12 strips of 8 wells) coated with a polyclonal antibody against human A1AT. Sealing Tapes: Each kit contains 3 precut, pressure sensitive sealing tapes that can be cut to fit the format of the individual assay. Human alpha-1-Antitrypsin Standard: Human A1AT in a buffered protein base (55 ng, lyophilized). Biotinylated Human alpha-1-Antitrypsin Antibody (60x): A 60-fold concentrated biotinylated polyclonal antibody against human A1AT (100 l). MIX Diluent Concentrate (10x): A 10-fold concentrated buffered protein base (30 ml). Wash Buffer Concentrate (20x): A 20-fold concentrated buffered surfactant (30 ml, 2 bottles). SP Conjugate (100x): A 100-fold concentrate (80 l). Chromogen Substrate (1x): A stabilized peroxidase chromogen substrate tetramethylbenzidine (8 ml). Stop Solution (1x): A 0.5 N hydrochloric acid solution to stop the chromogen substrate reaction (12 ml).
    Benötigtes Material
    Microplate reader capable of measuring absorbance at 405 nm. Pipettes (1-20 µL, 20-200 µL, and multiple channel). Deionized or distilled reagent grade water. Incubator (37 °C)
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  • Probenmenge
    50 μL
    Testdauer
    3 h
    Plattentyp
    Pre-coated
    Protokoll
    • Step 1. Add 50 μL of Standard or Sample per well. Incubate 2 hours.
    • Step 2. Wash, then add 50 μL of Biotinylated Antibody per well. Incubate 1 hour.
    • Step 3. Wash, then add 50 μL of SP Conjugate per well. Incubate 30 minutes.
    • Step 4. Wash, then add 50 μL of Chromogen Substrate per well. Incubate 8 minutes.
    • Step 5. Add 50 μL of Stop Solution per well. Read at 450 nm immediately.
    Aufbereitung der Reagenzien

    Freshly dilute all reagents and bring all reagents to room temperature before use. MIX Diluent Concentrate (10x): If crystals have formed in the concentrate, mix gently until the crystals have completely dissolved. Dilute the MIX Diluent Concentrate 10-fold with reagent grade water to produce a 1x solution. Store for up to 30 days at 2-8 °C. Human alpha-1-Antitrypsin Standard: Reconstitute the Human alpha-1- Antitrypsin Standard (55 ng) with 1.1 mL of MIX Diluent to generate a 50 ng/mL standard stock solution. Allow the vial to sit for 10 minutes with gentle agitation prior to making dilutions. Prepare duplicate or triplicate standard points by serially diluting from the standard stock solution (50 ng/mL) 4-fold with MIX Diluent to produce 12.5, 3.125, 0.781, and 0.195 ng/mL solutions. MIX Diluent serves as the zero standard (0 ng/mL). Any remaining stock solution should be stored at -20 °C and used within 30 days. Avoid repeated freeze-thaw cycles. Standard Point Dilution [A1AT] (ng/mL) P1 1 part Standard (50 ng/mL) 50 P2 1 part P1 + 3 parts MIX Diluent 12.5 P3 1 part P2 + 3 parts MIX Diluent 3.125 P4 1 part P3 + 3 parts MIX Diluent 0.781 P5 1 part P4 + 3 parts MIX Diluent 0.195 P6 MIX Diluent 0.0 5 Biotinylated Human alpha-1-Antitrypsin Antibody (60x): Spin down the antibody briefly and dilute the desired amount of the antibody 60-fold with MIX Diluent to produce a 1x solution. The undiluted antibody should be stored at -20 °C. Wash Buffer Concentrate (20x): If crystals have formed in the concentrate, mix gently until the crystals have completely dissolved. Dilute the Wash Buffer Concentrate 20-fold with reagent grade water to produce a 1x solution. SP Conjugate (100x): Spin down the SP Conjugate briefly and dilute the desired amount of the conjugate 100-fold with MIX Diluent to produce a 1x solution. The undiluted conjugate should be stored at -20 °C.

    Probennahme
    Plasma: Collect plasma using one-tenth volume of 0.1 M sodium citrate as an anticoagulant. Centrifuge samples at 3000 x g for 10 minutes and collect plasma. A 200000-fold sample dilution is suggested into MIX Diluent, however, user should determine optimal dilution factor depending on application needs. The undiluted samples can be stored at -20 °C or below for up to 3 months. Avoid repeated freeze-thaw cycles (EDTA or Heparin can also be used as an anticoagulant). Serum: Samples should be collected into a serum separator tube. After clot formation, centrifuge samples at 3000 x g for 10 minutes and remove serum. A 200000-fold sample dilution is suggested into MIX Diluent, however, user should determine optimal dilution factor depending on application needs. The undiluted samples can be stored at -20 °C or below for up to 3 months. Avoid repeated freeze-thaw cycles. Urine: Collect urine using sample pot. Centrifuge samples at 800 x g for 10 minutes. A 20-fold sample dilution is suggested into MIX Diluent or within the range of 2x - 200x, however, user should determine optimal dilution factor depending on application needs. The undiluted samples can be stored at -20 °C or below for up to 3 months. Avoid repeated freeze-thaw cycles. Saliva: Collect saliva using sample tube. Centrifuge samples at 800 x g for 10 minutes. A 400-fold sample dilution is suggested into MIX Diluent or within the range of 40x - 4000x, however, user should determine optimal dilution factor depending on application needs. The undiluted samples can be stored at -20 °C or below for up to 3 months. Avoid repeated freeze-thaw cycles. Milk: Collect milk using sample tube. Centrifuge samples at 800 x g for 10 minutes. A 2000-fold sample dilution is suggested into MIX Diluent, however, user should determine optimal dilution factor depending on application needs. The undiluted samples can be stored at -20 °C or below for up to 3 months. Avoid repeated freeze-thaw cycles. CSF: Collect cerebrospinal fluid (CSF) using sample pot. Centrifuge samples at 3000 x g for 10 minutes. A 4000-fold sample dilution is suggested into MIX Diluent, however, user should determine optimal dilution factor depending on application needs. The undiluted samples can be stored at -80 °C for up to 3 months. Avoid repeated freeze-thaw cycles. Cell Culture Supernatants: Centrifuge cell culture media at 3000 x g for 10 minutes at 4 °C to remove debris and collect supernatants. Samples can be stored at -20 °C or below. Avoid repeated freeze-thaw cycles.
    Testdurchführung

    Prepare all reagents, standard solutions, and samples as instructed. Bring all reagents to room temperature before use. The assay is performed at room temperature (20-25 °C). Remove excess microplate strips from the plate frame and return them immediately to the foil pouch with desiccants inside. Reseal the pouch securely to minimize exposure to water vapor and store in a vacuum desiccator. Add 50 l of Human alpha-1-Antitrypsin Standard or sample to each well. Gently tap plate to thoroughly coat the wells. Break any bubbles that may have formed. Cover wells with a sealing tape and incubate for 2 hours. Start the timer after the last addition. Wash five times with 200 l of Wash Buffer manually. Invert the plate each time and decant the contents, hit 4-5 times on absorbent material to completely remove the liquid. If using a machine, wash six times with 300 l of Wash Buffer and then invert the plate, decanting the contents, hit 4-5 times on absorbent material to completely remove the liquid. Add 50 l of Biotinylated Human alpha-1-Antitrypsin Antibody to each well. Gently tap plate to thoroughly coat the wells. Break any bubbles that may have formed. Cover wells with a sealing tape and incubate for 1 hour. Wash the microplate as described above. Add 50 l of SP Conjugate to each well. Gently tap plate to thoroughly coat the wells. Break any bubbles that may have formed. Cover wells with a sealing tape and incubate for 30 minutes. Turn on the microplate reader and set up the program in advance. Wash the microplate as described above. Add 50 l of Chromogen Substrate to each well. Gently tap plate to thoroughly coat the wells. Break any bubbles that may have formed. Incubate for 8 minutes or until the optimal blue color density develops. 6 Add 50 l of Stop Solution to each well. The color will change from blue to yellow. Gently tap plate to ensure thorough mixing. Break any bubbles that may have formed. Read the absorbance on a microplate reader at a wavelength of 450 nm immediately. If wavelength correction is available, subtract readings at 570 nm from those at 450 nm to correct optical imperfections. Otherwise, read the plate at 450 nm only. Please note that some unstable black particles may be generated at high concentration points after stopping the reaction for about 10 minutes, which will reduce the readings.

    Ergebnisberechnung
    • Calculate the mean value of the duplicate or triplicate readings for each standard and sample.
    • To generate a standard curve, plot the graph using the standard concentrations on the x-axis and the corresponding mean 450 nm absorbance (OD) on the y-axis. The best-fit line can be determined by regression analysis using log-log or four-parameter logistic curve-fit.
    • Determine the unknown sample concentration from the standard curve and multiply the value by the dilution factor.
    Testpräzision
    Intra-assay and inter-assay coefficients of variation were 4.1% and 7.1% respectively.
    Beschränkungen
    Nur für Forschungszwecke einsetzbar
  • Handhabung
    This product is for Research Use Only and is not intended for use in diagnostic procedures. Prepare all reagents (diluent buffer, wash buffer, standard, biotinylated antibody, and SP conjugate) as instructed, prior to running the assay. Prepare all samples prior to running the assay. The dilution factors for the samples are suggested in this insert. However, the user should determine the optimal dilution factor. Spin down the SP conjugate vial and the biotinylated antibody vial before opening and using contents. The Stop Solution is an acidic solution. The kit should not be used beyond the expiration date. 2
    Lagerung
    4 °C/-20 °C
    Informationen zur Lagerung
    Upon arrival, immediately store components of the kit at recommended temperatures up to the expiration date. Store SP Conjugate and Biotinylated Antibody at -20°C. Store Microplate, Diluent Concentrate (10x), Wash Buffer, Stop Solution, and Chromogen Substrate at 2-8°C. Unused microplate wells may be returned to the foil pouch with the desiccant packs and resealed. May be stored for up to 30 days in a vacuum desiccator. Diluent (1x) may be stored for up to 30 days at 2-8°C. Store Standard at 2-8°C before reconstituting with Diluent and at -20°C after reconstituting with Diluent.
  • Target Alle SERPINA1 ELISA Kits anzeigen
    SERPINA1 (serpin Peptidase Inhibitor, Clade A (Alpha-1 Antiproteinase, Antitrypsin), Member 1 (SERPINA1))
    Andere Bezeichnung
    Alpha-1-Antitrypsin (A1AT) (SERPINA1 Produkte)
    Synonyme
    AACT ELISA Kit, ACT ELISA Kit, GIG25 ELISA Kit, A1AT ELISA Kit, PI ELISA Kit, alpha-1-PI ELISA Kit, A1A ELISA Kit, AAT ELISA Kit, PI1 ELISA Kit, PRO2275 ELISA Kit, alpha1AT ELISA Kit, Pi ELISA Kit, Spi1 ELISA Kit, MGC64429 ELISA Kit, serpina1d ELISA Kit, Aps ELISA Kit, SERPINA3-5 ELISA Kit, serpin family A member 3 ELISA Kit, alpha-1-antiproteinase F precursor ELISA Kit, serpin family A member 1 ELISA Kit, alpha-1-antitrypsin ELISA Kit, calumenin ELISA Kit, serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 1 S homeolog ELISA Kit, serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 1 ELISA Kit, serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 5 ELISA Kit, SH2B adaptor protein 2 ELISA Kit, serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 3 ELISA Kit, serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, antitrypsin), member 1 L homeolog ELISA Kit, SERPINA3 ELISA Kit, LOC100008973 ELISA Kit, SERPINA1 ELISA Kit, LOC100328621 ELISA Kit, CALU ELISA Kit, Serpina1 ELISA Kit, serpina1.S ELISA Kit, serpina1 ELISA Kit, SPIA5 ELISA Kit, Sh2b2 ELISA Kit, Calu ELISA Kit, serpina1.L ELISA Kit
    Hintergrund
    Alpha-1-antitrypsin (A1AT) is a protein that protects the lungs. The liver usually makes the protein and releases it into the bloodstream. A1AT is a major protease inhibitor that controls tissue degradation. A reduction of A1AT levels can cause a change in collagen metabolism (1). A1AT inhibits neutrophil elastase release into the lungs during inflammatory states (2).
    Gen-ID
    5265
    UniProt
    P01009
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