Neuroplastin (NPTN) BioAssay(TM) ELISA Kit (Human)

Product Overview

• Article Name: Neuroplastin (NPTN) BioAssay(TM) ELISA Kit (Human)
• Biozol Catalog Number: USB-152878
• Supplier Catalog Number: 152878
• Alternative Catalog Number: USB-152878-96
• Manufacturer: US Biological
• Category: Kits/Assays

Product Description

The Human Neuroplastin (NPTN) ELISA Kit is a sandwich enzyme immunoassay for the in vitro quantitative measurement of NPTN in human tissue homogenates and other biological fluids. Detection Range: 0.156-10ng/ml Sensitivity: <0.052ng/ml Precision: Intra-Assay: CV<10% Inter-Assay: CV<12% Test Principle: The microtiter plate provided in this kit has been pre-coated with an antibody specific to NPTN. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody specific to NPTN. Next, Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After TMB substrate solution is added, only those wells that contain NPTN, biotin-conjugated antibody and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of sulfuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450nm 10nm. The concentration of NPTN in the sample is then determined by comparing the O.D. of the sample to the standard curve. Kit Components: *152878A: Microtiter Plate, 1x96 wells, Pre-coated, ready to use *152878B: Standard, 2x1vial 152878C: Standard Diluent, 1x20ml *152878D: Detection Reagent A, 1x120ul *152878E: Detection Reagent B, 1x120ul 152878F: Assay Diluent A, 1x12ml 152878G: Assay Diluent B, 1x12ml 152878H: TMB Substrate, 1x9ml 152878K: Stop Solution, 1x6ml 152878L: Wash Buffer, 30X, 1x20ml Precaution: The Stop Solution (152878K) 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 *152878A, *152878B, *152878D and *152878E 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: 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 2500*g. The supernatant was removed and assayed immediately or aliquoted and stored at -20C or lower. Other Biological Fluids: Centrifuge samples for 20 minutes at 1000*g. Remove particulates and assay immediately or store samples in aliquots at -20C or -80C. Avoid repeated freeze/thaw cycles. Note: 1. Samples to be used within 5 days may be stored at 4C, otherwise samples must be stored at -20C (1 month) or -80C (2 months) to avoid loss of bioactivity and contamination. 2. When performing the assay, bring samples to RT. Assay Procedure Summary: 1. Prepare all reagents, samples and standards. 2. Add 100ul standard or sample to each well. Incubate 2 hours at 37C. 3. Aspirate and add 100ul prepared Detection Reagent A. Incubate 1 hour at 37C. 4. Aspirate and wash 3 times. 5. Add 100ul prepared Detection Reagent B. Incubate 30 minutes at 37C. 6. Aspirate and wash 5 times. 7. Add 90ul TMB Substrate. Incubate 15-25 minutes at 37C. 8. Add 50ul Stop Solution. Read at 450nm immediately.