Human ubiquitin protein (Ub) elisa kit instruction manual

**Human Ubiquitin Protein (Ub) ELISA Kit – User Manual** This ELISA kit is designed for the quantitative determination of human ubiquitin protein (Ub) in various biological samples such as serum, plasma, urine, cell culture supernatants, and tissue homogenates. The kit utilizes a sandwich ELISA method, where specific antibodies are used to capture and detect Ub molecules in the sample. **Kit Specifications:** - **Configuration:** 48-well or 96-well format - **Standard Dilution:** 1.5 mL × 1 vial - **Enzyme Standard Reagent:** 3 mL × 1 vial (for 48-well), 6 mL × 1 vial (for 96-well) - **Storage Conditions:** 2–8°C - **Validity Period:** 6 months from date of receipt **Kit Components:** - Sealing Film: 2 pieces (48) / 2 pieces (96) - Standard: 0.5 mL × 1 vial, 2700 ng/L - Enzyme Standard: 1×48 / 1×96 - Sample Diluent: 3 mL × 1 vial (48) / 6 mL × 1 vial (96) - Developer A: 3 mL × 1 vial (48) / 6 mL × 1 vial (96) - Developer B: 3 mL × 1 vial (48) / 6 mL × 1 vial (96) - Wash Buffer: 3 mL × 1 vial (48) / 6 mL × 1 vial (96) - Concentrated Wash Solution: 20 mL × 20 times (48) / 20 mL × 30 times (96) **Principle of Operation:** The assay is based on a double-antibody sandwich technique. The microtiter plate is pre-coated with a monoclonal anti-Ub antibody. After incubation with the sample, Ub binds to the immobilized antibody. A horseradish peroxidase (HRP)-conjugated secondary antibody is then added, forming an immune complex. TMB substrate is used for color development, and the intensity of the color is directly proportional to the concentration of Ub in the sample. Absorbance is measured at 450 nm, and the concentration is calculated using a standard curve. **Sample Preparation Guidelines:** - **Serum/Plasma:** Centrifuge at 2000–3000 rpm for 20 minutes after clotting or anticoagulant addition. - **Urine:** Centrifuge at 2000–3000 rpm for 20 minutes. - **Cell Culture Supernatant:** Centrifuge to remove debris; for intracellular components, lyse cells by freezing and thawing before centrifugation. - **Tissue Homogenate:** Homogenize in PBS, centrifuge, and collect the supernatant. **Procedure Summary:** 1. Prepare standard dilutions and load into designated wells. 2. Add sample diluent and test samples to the microplate. 3. Incubate at 37°C for 30 minutes. 4. Wash the plate 5 times with wash buffer. 5. Add enzyme-labeled reagent and incubate again. 6. Add TMB substrate and develop color for 15 minutes. 7. Stop the reaction with stop solution. 8. Measure absorbance at 450 nm using a microplate reader. 9. Calculate concentrations using the standard curve. **Important Notes:** - Allow the kit to equilibrate at room temperature before use. - Avoid repeated freeze-thaw cycles of samples. - Do not mix reagents from different batches. - Use a new sealing film for each experiment to prevent contamination. - Handle all reagents with care and dispose of waste properly. **Performance Characteristics:** - **Linear Range:** 0.2 IU/L – 6 IU/L - **Correlation Coefficient (R²):** ≥ 0.95 - **Intra-assay CV:** < 9% - **Inter-assay CV:** < 11% **Service Commitment:** We offer free technical support during working hours and can assist with sample testing upon request. Delivery is typically within 5–7 business days after payment. **Storage & Handling:** Store the kit at 2–8°C. Avoid exposure to light and moisture. Always follow the instructions carefully for accurate results. This kit is intended for research use only and should not be used for diagnostic purposes.

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