Hemoglobin (HB) BioAssay(TM) ELISA Kit (Porcine) is a competitive inhibition enzyme immunoassay for the in vitro quantitative measurement of HB in porcine serum, plasma and erythrocyte lysates. Detection Range: 3.70-300ug/ml Sensitivity: 1.42ug/ml Intra-Assay CV: <10% Inter-Assay CV: <12% Test Principle: This assay employs the competitive inhibition enzyme immunoassay technique. A monoclonal antibody specific for porcine HB has been pre-coated onto a microplate. A competitive inhibition reaction occurs between biotin-labeled porcine HB and unlabeled porcine HB (standards or samples) with the pre-coated antibody specific for porcine 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 the color developed is inversely proportional to the concentration of HB in the sample. Kit Components: *025603A: Microtiter Strips, 1x96 wells, Pre-coated, ready to use *025603B: Standard, 2x1vial 025603C: Standard Diluent, 1x20ml *025603D: Detection Reagent A, 1x120ul *025603E: Detection Reagent B, 1x120ul 025603F: Assay Diluent A, 1x12ml 025603G: Assay Diluent B, 1x12ml 025603H: TMB Substrate, 1x9ml 025603K: Stop Solution, 1x6ml 025603L: Wash Buffer, 30X, 1x20ml Precaution: The Stop Solution (025603K) included for use with this kit is an acid solution. Wear eye, hand, face, and clothing protection when using this material. Storage and Stability: Store *025603A, *025603B, *025603D and *025603E 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. Assay Procedure Summary: 1. Prepare all reagents, samples and standards. 2. Add 50ul standard or sample to each well, then add 50ul prepared Detection Reagent A immediately. Shake and mix. Incubate 1 hour at 37C. 3. Aspirate and wash 3 times. 4. Add 100ul prepared Detection Reagent B. Incubate 30 minutes at 37C. 5. Aspirate and wash 5 times. 6. Add 90ul Substrate Solution. Incubate 15-25 minutes at 37C. 7. Add 50ul Stop Solution. Read at 450 nm immediately.
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