Mouse interleukin 18 (IL-18) elisa kit instruction manual

**Mouse IL-18 (Interleukin-18) ELISA Kit – User Manual** This ELISA kit is designed for the quantitative determination of Mouse Interleukin-18 (IL-18) in biological samples such as serum, plasma, urine, cell culture supernatants, and tissue homogenates. The kit utilizes a sandwich ELISA method, where a monoclonal antibody specific to IL-18 is pre-coated on the microplate. After incubation with the sample, a horseradish peroxidase (HRP)-labeled secondary antibody binds to the captured IL-18, forming a complex. A TMB substrate is then added, and the reaction is stopped with an acidic solution, resulting in a color change that correlates with the concentration of IL-18 in the sample. Absorbance is measured at 450 nm using a microplate reader, and concentrations are calculated using a standard curve. **Kit Specifications:** - **Configuration:** 48-well or 96-well format - **Standard Dilution:** 1.5 mL × 1 vial - **Enzyme Standard Reagent:** 3 mL × 1 vial (48-well), 6 mL × 1 vial (96-well) - **Storage Conditions:** 2–8°C - **Validity:** 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) - Sample diluent: 3 mL × 1 vial (48) / 6 mL × 1 vial (96) - HRP-conjugated antibody: 3 mL × 1 vial (48) / 6 mL × 1 vial (96) - TMB substrate A & B: 3 mL × 1 vial each - Washing buffer: 3 mL × 1 vial (48) / 6 mL × 1 vial (96) - Concentrated washing solution: 20 mL × 20 times (48) / 20 mL × 30 times (96) **Sample Preparation Instructions:** - **Serum/Plasma:** Centrifuge at 2000–3000 rpm for 20 minutes after clotting (serum) or anticoagulant addition (plasma). - **Urine:** Centrifuge at 2000–3000 rpm for 20 minutes. - **Cell Culture Supernatant:** Centrifuge at 2000–3000 rpm for 20 minutes. For intracellular components, lyse cells via freeze-thaw cycles before centrifugation. - **Tissue Homogenate:** Weigh the tissue, homogenize in PBS (pH 7.4), centrifuge, and collect supernatant. **Operating Procedure:** 1. Prepare a serial dilution of the IL-18 standard (1800 ng/L to 150 ng/L). 2. Add 40 µL of sample diluent and 10 µL of sample to each well (final dilution: 5×). 3. Incubate at 37°C for 30 minutes. 4. Wash the plate 5 times with diluted washing buffer. 5. Add 50 µL of HRP-labeled antibody to each well (except blank wells). 6. Incubate again at 37°C for 30 minutes. 7. Wash the plate again. 8. Add 50 µL of TMB substrate A and B, incubate for 15 minutes at 37°C. 9. Stop the reaction by adding 50 µL of stop solution. 10. Measure OD450 immediately within 15 minutes. **Data Analysis:** Plot the standard curve using the OD values vs. concentrations. Calculate the sample concentration using linear regression or interpolate from the curve. Multiply by the dilution factor to obtain the actual concentration. **Notes:** - Allow all reagents to reach room temperature before use. - Avoid repeated freezing and thawing of samples. - Do not mix reagents from different batches. - Keep substrates away from light. - All waste should be treated as biohazardous material. **Performance:** - 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 to ensure accurate results. **Storage:** Store the kit at 2–8°C. Do not freeze. This manual is intended for research purposes only. Always follow the instructions carefully to ensure reliable results.

Shoe Cabinet

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