Hemoglobin (HB) BioAssay(TM) ELISA Kit (Human)

Produktübersicht

• Artikelname: Hemoglobin (HB) BioAssay(TM) ELISA Kit (Human)
• Artikelnummer: USB-025597
• Hersteller Artikelnummer: 025597
• Alternativnummer: USB-025597-96
• Hersteller: US Biological
• Kategorie: Kits/Assays

Produktbeschreibung

The Human Hemoglobin (HB) ELISA Kit is a competitive inhibition immunoassay for the in vitro quantitative measurement of HB in human serum, plasma, tissue homogenates, erythrocyte lysates, cell culture supernatants and other biological fluids. Detection Range: 1.23-100ug/ml Sensitivity: <0.51ug/ml Precision: Intra-Assay CV: <10% Inter-Assay CV: <12% Test Principle: This assay employs the competitive inhibition enzyme immunoassay technique. A monoclonal antibody specific for HB has been pre-coated onto a microplate. A competitive inhibition reaction occurs between biotin-labeled HB and unlabeled HB (standards or samples) for limited binding sites on the pre-coated antibody specific for HB. After incubation the unbound conjugate is washed off. Next, avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. The amount of bound HRP conjugate is inversely proportional to the concentration of HB in the sample. After addition of the substrate solution, the intensity of color developed is inversely proportional to the concentration of HB in the sample. Kit Components: *025597A: Microtiter Plate, 96 wells, Pre-coated, ready to use *025597B: Standard, 2x1vial 025597C: Standard Diluent, 1x20ml *025597D: Detection Reagent A, 1x120ul *025597E: Detection Reagent B, 1x120ul 025597F: Assay Diluent A, 1x12ml 025597G: Assay Diluent B, 1x12ml 025597H: TMB Substrate, 1x9ml 025597K: Stop Solution, 1x6ml 025597L: Wash Buffer, 30X, 1x20ml Precaution: The Stop Solution (025597K) suggested for use with this kit is an acid solution. Wear eye, hand, face, and clothing protection when using this material. Storage and Stability: Store *025597A, *025597B, *025597D and *025597E at -20C. Store all the other components at 4C. Unused kit is stable for 6 months. Once kit components are opened, it is highly recommended to use remaining reagents within 1 month provided this is within the expiration date of the kit. For maximum recovery of product, centrifuge the original vials after thawing and prior to removing the cap. Materials Required But Not Supplied: 1. Microplate reader with 450 10nm filter. 2. Precision single or multi-channel pipettes and disposable tips. 3. Eppendorf Tubes for diluting samples. 4. Deionized or distilled water. 5. Absorbent paper for blotting the microtiter plate. 6. Container for Wash Solution Sample Preparation and Storage: Serum: Centrifuge whole blood for 20 minutes at approximately 1000xg. Remove serum and assay immediately or store samples in aliquots at -20C or -80C. Avoid repeated freeze/thaw cycles. Plasma: Collect plasma using EDTA as anticoagulant. Centrifuge samples for 15 minutes at 1000xg at 2-8C within 30 minutes of collection. Remove plasma and assay immediately or store samples in aliquots at -20C or -80C for later use. Avoid repeated freeze/thaw cycles. Tissue Homogenates: The preparation of tissue homogenates will vary depending upon tissue type. For this assay, tissues were rinsed thoroughly in ice-cold PBS (0.02M pH 7.0-7.2) to remove excess blood and weighed before homogenization. The tissues were minced to small pieces and homogenized in 5-10ml of PBS with a glass homogenizer on ice (Micro Tissue Grinders works too). The resulting suspension was sonicated with an ultrasonic cell disrupter or subjected to two freeze-thaw cycles to further break the cell membranes. After that, the homogenates were centrifuged for 5 minutes at 5000*g. The supernatant was removed and assayed immediately or aliquoted and stored at -20C or lower. Note: If free HB is detected in non-hemolyzed serum and plasma, samples require about 10-fold dilution. A suggested 10-fold dilution is 20ul sample + 180ul PBS. Samples should be diluted using 0.01M PBS, pH 7.0-7.2. Erythrocyte Lysates: 1. Centrifuge whole blood for 20 minutes at approximately 1000xg, remove the supernatant and collect the cells. 2. Wash cells three times with PBS (0.01M, pH 7.0 - 7.2). 3. Resuspend cells in ice-cold PBS and fr