Horse β-endorphin (β-EP) elisa kit instruction manual

**Horse β-Endorphin (β-EP) ELISA Kit – Instructions for Use** **Kit Specifications:** This kit is available in 48-well or 96-well configurations. It includes a standard dilution of 1.5 ml × 1 bottle, enzyme standard reagent (3 ml × 1 bottle for 48-well; 6 ml × 1 bottle for 96-well). The kit is intended for research use only. **Calculation Method:** To determine the concentration of β-EP in the sample, plot the standard concentrations on the x-axis and the corresponding OD values on the y-axis to generate a standard curve. Alternatively, calculate the linear regression equation using the standard concentrations and their OD values, then substitute the sample’s OD value into the equation. Multiply the result by the dilution factor to obtain the actual sample concentration. **Kit Components:** - Sealing film: 2 pieces (48-well) / 2 pieces (96-well) - Standard: 0.5 ml × 1 bottle, 2700 ng/L - Enzyme standard: 1×48 / 1×96 - Sample diluent: 3 ml × 1 bottle / 6 ml × 1 bottle - Developer A: 3 ml × 1 bottle / 6 ml × 1 bottle - Chromogen B: 3 ml × 1 bottle / 6 ml × 1 bottle - Stop solution: 3 ml × 1 bottle / 6 ml × 1 bottle - Concentrated wash solution: (20 ml × 20 times) × 1 bottle / (20 ml × 30 times) × 1 bottle **Storage Conditions:** Store all components at 2–8°C. The shelf life is 6 months from the date of receipt. **Principle of the Assay:** The kit uses a double-antibody sandwich ELISA method. Microtiter plates are pre-coated with anti-β-EP antibodies. After adding the samples and standards, β-EP binds to the immobilized antibody. HRP-conjugated secondary antibody is then added, forming an immune complex. TMB substrate is used for color development, and the reaction is stopped with a stop solution. The absorbance at 450 nm is measured, and the β-EP concentration is determined using a standard curve. **Purpose:** This ELISA kit is designed to quantify β-endorphin (β-EP) levels in horse serum, plasma, urine, cell culture supernatants, tissue homogenates, and other biological fluids. **Sample Preparation Guidelines:** - **Serum:** Allow blood to clot at room temperature for 10–20 minutes, centrifuge at 2000–3000 rpm for 20 minutes. - **Plasma:** Use EDTA or sodium citrate as anticoagulant, mix for 10–20 minutes, centrifuge similarly. - **Urine:** Centrifuge for 20 minutes at 2000–3000 rpm. - **Cell Culture Supernatant:** Centrifuge after collection; for intracellular components, lyse cells via freeze-thaw cycles before centrifugation. - **Tissue:** Homogenize in PBS, centrifuge, and collect supernatant. Store at 2–8°C after thawing. **Notes for Operation:** - Equilibrate the kit at room temperature for 15–30 minutes before use. - Avoid repeated freezing and thawing of samples. - Do not use samples containing NaN₃, as it may inhibit HRP activity. - Always prepare a standard curve and perform duplicate measurements for accuracy. - Handle all reagents carefully and follow the protocol precisely. - Discard all waste materials as biohazardous. - Do not mix components from different batches. **Performance Characteristics:** - Correlation coefficient (R) ≥ 0.95 - Intra-assay CV < 9%, Inter-assay CV < 11% - Detection range: 0.2 IU/L – 6 IU/L **Service Commitment:** We offer free technical support during working hours and can assist with sample testing to ensure accurate results. Delivery time is within the agreed period. **Usage Tips:** - Ensure all pipetting steps are performed accurately and quickly (within 5 minutes per step). - For large numbers of samples, use an automated pipette. - Keep the substrate away from light during incubation. - Read the plate within 15 minutes after adding the stop solution. This kit provides a reliable and efficient method for measuring β-EP levels in equine samples, ensuring high sensitivity and reproducibility.

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