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Hemopexin ELISA 试剂盒

HPX 适用: 人 Colorimetric Sandwich ELISA Cell Culture Supernatant, Plasma
产品编号 ABIN612796
发货至: 中国
  • 抗原 See all Hemopexin (HPX) ELISA试剂盒
    Hemopexin (HPX)
    适用
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    检测方法
    Colorimetric
    实验类型
    Sandwich ELISA
    最低检测浓度
    4 ng/mL
    应用范围
    ELISA
    原理
    The AssayMax Human Hemopexin ELISA kit is designed for detection of human and murine Hemopexin in plasma, serum and cell culture supernatants
    品牌
    AssayMax
    样品类型
    Plasma, Cell Culture Supernatant
    Analytical Method
    Quantitative
    组件
    Hemopexin Microplate: A 96-well polystyrene microplate (12 strips of 8 wells) coated with a polyclonal antibody against human hemopexin. Sealing Tapes: Each kit contains 3 pre-cut, pressure-sensitive sealing tapes, which can be cut to fit the format of the individual assay. Hemopexin Standard: Human Hemopexin in a buffered protein base (800 ng, lyophilized). Biotinylated Hemopexin Antibody (100x): A 100-fold concentrated biotinylated polyclonal antibody against hemopexin (90µl). 1 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). Streptavidin-Peroxidase Conjugate (SP Conjugate): A 100-fold concentrate (90µl). Chromogen Substrate: A ready-to-use stabilized peroxidase chromogen substrate tetramethylbenzidine (8 ml). Stop Solution: A 0.5 N hydrochloric acid to stop the chromogen substrate reaction (12 ml).
    试剂未包括
    Microplate reader capable of measuring absorbance at 450 nm. Pipettes (1-20 µL, 20-200 µL, 200-1000µLand multiple channel). Deionized or distilled reagent grade water.
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  • 样本量
    50 μL
    实验时间
    3.5 h
    板类型
    Pre-coated
    实验流程
    This assay employs a quantitative sandwich enzyme immunoassay technique that measures hemopexin in 3.5 hours. A polyclonal antibody specific for hemopexin has been pre-coated onto a microplate. Hemopexin in standards and samples is sandwiched by the immobilized antibody and biotinylated polyclonal antibody specific for hemopexin, which is recognized by a streptavidin-peroxidase conjugate. All unbound material is then washed away and a peroxidase enzyme substrate is added. The color development is stopped and the intensity of the color is measured.
    试剂准备

    Freshly dilute all reagents and bring all reagents to room temperature before use. If crystals have formed in the concentrate, mix gently until the crystals have completely dissolved. MIx Diluent Concentrate (10x): Dilute the MIx Diluent 1:10 with reagent grade water. Store for up to 1 month at 2-8°C. Hemopexin Standard: Reconstitute the 800 ng of human hemopexin Standard with 2 ml of MIx Diluent to generate a stock solution of 400 ng/ml. Allow the standard to sit for 10 minutes with gentle agitation prior to making dilutions. Prepare triplicate standard points by serially diluting the Standard solution (400 ng/ml) 1:2 with MIx Diluent to produce 200, 100, 50, 25, 12.5 and 6.25 ng/ml. Sample Diluent serves as the zero standard (0 ng/ml). Any remaining solution should be frozen at -20°C. Standard Point Dilution [Hemopexin] (ng/ml) P1 1 part Standard (400 ng/ml) 400.00 P2 1 part P1 + 1 part MIx Diluent 200.00 P3 1 part P2 + 1 part MIx Diluent 100.00 P4 1 part P3 + 1 part MIx Diluent 50.00 P5 1 part P4 + 1 part MIx Diluent 25.00 P6 1 part P5 + 1 part MIx Diluent 12.50 P7 1 part P6 + 1 part MIx Diluent 6.25 P8 MIx Diluent 0.00 Biotinylated Hemopexin Antibody (100x): Spin down the SP Conjugate briefly and dilute the desired amount of the antibody 1:100 with MIx Diluent. Any remaining solution should be frozen at -20°C. Wash Buffer Concentrate (20x): Dilute the Wash Buffer Concentrate 1:20 with reagent grade water. SP Conjugate (100x): Spin down the SP Conjugate briefly and dilute the desired amount of the conjugate 1:100 with MIx Diluent. Any remaining solution should be frozen at -20°C.

    样品收集
    Cell Culture Supernatants: Centrifuge cell culture media at 2000 x g for 10 minutes to remove debris. Collect supernatants and assay. Dilute samples 1:10 into MIx Diluent. Store samples at -20°C or below. Avoid repeated freeze-thaw cycles. Urine: Collect urine using sample pot. Centrifuge samples at 600 x g for 10 minutes and assay. Dilute samples 1:10 into MIx Diluent. Store samples at -20°C or below for up to 3 months. Avoid repeated freeze-thaw cycles.
    实验流程

    Prepare all reagents, working standards and samples as instructed. Bring all reagents to room temperature before use. The assay is performed at room temperature (20 - 30 °C). Remove excess microplate strips from the plate frame and return them immediately to the foil pouch with desiccant inside. Reseal the pouch securely to minimize exposure to water vapor and store in a vacuum desiccator. Add 50 µL of Standard or sample per well. Cover wells with a sealing tape and incubate for two hours. Start the timer after the last sample addition. Wash five times with 200 µL of Wash Buffer. Invert the plate and decant the contents, and hit it 4-5 times on absorbent paper towel to complete remove liquid at each step. Add 50 µL of Biotinylated Hemopexin Antibody to each well and incubate for one hour. Wash five times with 200 µL of Wash Buffer. Add 50 µL of Streptavidin-Peroxidase Conjugate per well and incubate for 30 minutes. Turn on the microplate reader and set up the program in advance. Wash five times with 200 µL of Wash Buffer. Add 50 µL of Chromogen Substrate per well and incubate for about 10 minutes or till the optimal blue color density develops. Gently tap plate to ensure thorough mixing and break the bubbles in the well with pipette tip. Add 50 µL of Stop Solution to each well. The color will change from blue to yellow. Read the absorbance on a microplate reader at a wavelength of 450 nm immediately. 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.

    结果分析

    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 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. Standard Curve The curve is provided for illustration only. A standard curve should be generated each time the assay is performed.

    实验精密度
    Intra-assay and inter-assay coefficients of variation were 4.8 % and 7.5% respectively.
    限制
    仅限研究用
  • 注意事项
    The kit should not be used beyond the expiration date.
    储存条件
    4 °C/-20 °C
    储存方法
    Store kit at 2-8°C or -20°C upon arrival up to the expiration date. Opened MIx Diluent may be stored for up to 1 month at 2-8°C. Store reconstituted reagents at -20°C or below. Opened unused strip wells may return to the foil pouch with the desiccant pack, reseal along zip-seal. May be stored for up to 1 month in a vacuum desiccator.
  • Pohl, Danz, Gross, John, Urban, Patzer, Habbig, Feldkötter, Witzke, Walther, Rhode: "Diagnosis of Alport syndrome-search for proteomic biomarkers in body fluids." in: Pediatric nephrology (Berlin, Germany), (2013) (PubMed).

  • 抗原 See all Hemopexin (HPX) ELISA试剂盒
    Hemopexin (HPX)
    别名
    Hemopexin (HPX 产品)
    别名
    HPX ELISA Kit, HX ELISA Kit, Hpxn ELISA Kit, hx ELISA Kit, wu:fi17f11 ELISA Kit, wu:fi19a10 ELISA Kit, hemopexin ELISA Kit, HPX ELISA Kit, Mmar10_1124 ELISA Kit, Hpx ELISA Kit, hpx ELISA Kit
    背景
    Hemopexin is a heme-binding plasma glycoprotein which, after haptoglobin, forms the second line of defense against hemoglobin-mediated oxidative damage during intravascular hemolysis. A decrease in plasma hemopexin concentration reflects a recent release of heme compounds in the extracellular compartment. Heme-hemopexin complexes are delivered to hepatocytes by receptor-mediated endocytosis after which hemopexin is recycled to the circulation. Studies indicated that increased hemopexin level associate with minimal change disease (MCD) , sporadic Alzheimer's disease (AD) , heavy-drinking chronic alcoholics , hemolytic anemias, chronic neuromuscular diseases and acute intermittent porphyria.
    途径
    Transition Metal Ion Homeostasis, Regulation of Leukocyte Mediated Immunity, Positive Regulation of Immune Effector Process, Production of Molecular Mediator of Immune Response
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