Human anti-alveolar basement membrane antibody (ABM-Ab) elisa kit instruction manual

**Human Anti-Alveolar Basement Membrane Antibody (ABM-Ab) ELISA Kit Instruction Manual** **Kit Specifications:** This ELISA kit is available in a 48-well or 96-well configuration. It includes the following components: - Standard: 1.5 ml × 1 vial - Enzyme Standard Reagent: 3 ml × 1 vial (for 48-well) / 6 ml × 1 vial (for 96-well) - Sealing Film: 2 pieces (for 48-well) / 2 pieces (for 96-well) - Microplate: 1×48 / 1×96 - Sample Diluent: 3 ml × 1 vial (for 48-well) / 6 ml × 1 vial (for 96-well) - Developer A: 3 ml × 1 vial (for 48-well) / 6 ml × 1 vial (for 96-well) - Chromogen B: 3 ml × 1 vial (for 48-well) / 6 ml × 1 vial (for 96-well) - Stop Solution: 3 ml × 1 vial (for 48-well) / 6 ml × 1 vial (for 96-well) - Concentrated Washing Solution: 20 ml × 20 times (for 48-well) / 20 ml × 30 times (for 96-well) **Storage Conditions:** All reagents should be stored at 2–8°C, except for Chromogen B, which should be stored at -20°C. The kit has a shelf life of 6 months from the date of manufacture. **Purpose:** This ELISA kit is designed to quantitatively determine the concentration of Human Anti-Alveolar Basement Membrane Antibody (ABM-Ab) in human serum, plasma, urine, and other biological fluids. **Principle of Operation:** The kit utilizes a sandwich ELISA method. A solid-phase antibody specific to ABM-Ab is coated on microtiter plates. After incubation with the sample, a horseradish peroxidase (HRP)-labeled secondary antibody binds to the complex. TMB substrate is then added, producing a color change that is proportional to the amount of ABM-Ab present. The reaction is terminated with a stop solution, and the absorbance is measured at 450 nm using a microplate reader. A standard curve is plotted, and the sample concentration is determined accordingly. **Sample Preparation:** - **Serum:** Allow blood to clot at room temperature, centrifuge at 2000–3000 rpm for 10–20 minutes, and collect the supernatant. - **Plasma:** Use EDTA or sodium citrate as anticoagulant, mix well, centrifuge, and collect supernatant. - **Urine:** Centrifuge at 2000–3000 rpm for 20 minutes, collect the supernatant. - **Cell Culture Supernatant:** Centrifuge and collect supernatant; for intracellular components, lyse cells by freezing and thawing. - **Tissue Samples:** Homogenize in PBS, centrifuge, and collect the supernatant. **Note:** Avoid repeated freeze-thaw cycles and store samples at -20°C if not tested immediately. Do not use samples containing NaN3, as it may inhibit HRP activity. **Operation Steps:** 1. Prepare standard dilutions and add to designated wells. 2. Add sample diluent and test samples to the appropriate wells. 3. Seal the plate and incubate at 37°C for 30 minutes. 4. Wash the plate 5 times with diluted washing solution. 5. Add enzyme-labeled reagent and incubate again. 6. Add TMB substrate and develop color for 15 minutes at 37°C. 7. Stop the reaction with stop solution and measure OD at 450 nm within 15 minutes. **Important Notes:** - Allow all reagents to reach room temperature before use. - Ensure accurate pipetting and avoid cross-contamination. - Always prepare a standard curve and run duplicates for better accuracy. - Store unused strips in sealed bags after opening. - Keep substrates away from light. - Follow instructions carefully and validate results with a microplate reader. - Dispose of all waste as biohazardous material. **Performance:** - Linear regression correlation coefficient (R) ≥ 0.95 - Intra-assay and inter-assay variation < 9% and < 11%, respectively - Detection range: 0.2 IU/L – 6 IU/L **Service Commitment:** - Free technical support during working hours - Sample testing services available upon request - Prompt delivery after payment - English manual takes precedence in case of discrepancies This kit is intended for research use only and not for diagnostic purposes.

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