Human aspartate aminotransferase/aspartate aminotransferase (GOT/GPT) elisa kit instruction manual

**Human Aspartate Aminotransferase (GOT/GPT) ELISA Kit – Instructions for Use** **Kit Specifications:** This ELISA kit is available in 48-well or 96-well configurations. The standard dilution is 1.5 mL × 1 vial. The enzyme standard reagent is 3 mL × 1 vial (for 48-well) or 6 mL × 1 vial (for 96-well). This reagent is intended for research use only. **Standard Curve Preparation:** To determine the concentration of the sample, plot the standard concentrations on the x-axis and their corresponding OD values on the y-axis. Draw a standard curve and estimate the sample concentration based on its OD value. Alternatively, calculate the linear regression equation from the standard curve data and substitute the sample’s OD into the equation to obtain the actual concentration after multiplying by the dilution factor. **Kit Composition:** - Sealing film: 2 pieces (48-well) / 2 pieces (96-well) - Standard: 0.5 mL × 1 vial (2700 ng/L) - Enzyme standard: 3 mL × 1 vial (48-well) / 6 mL × 1 vial (96-well) - Developer A: 3 mL × 1 vial / 6 mL × 1 vial - Chromogen B: 3 mL × 1 vial / 6 mL × 1 vial - Wash solution: 3 mL × 1 vial / 6 mL × 1 vial - Concentrated wash solution: 20 mL × 20 times / 20 mL × 30 times - All reagents should be stored at 2–8°C. **Principle of Operation:** This kit uses the sandwich ELISA method to detect human aspartate aminotransferase (GOT/GPT). The microplate is pre-coated with purified anti-GOT/GPT antibodies. After adding the sample, the target antigen binds to the immobilized antibody. HRP-labeled secondary antibody then binds to the complex, forming an antibody-antigen-enzyme conjugate. TMB substrate is added, producing a color change that is proportional to the antigen concentration. The reaction is stopped with an acidic solution, and the absorbance is measured at 450 nm using a microplate reader. **Purpose:** The kit is designed to quantify GOT/GPT levels in human serum, plasma, urine, cell culture supernatants, tissue homogenates, and other biological fluids. **Storage & Shelf Life:** Store the kit at 2–8°C. The shelf life is 6 months from the date of receipt. **Sample Preparation:** 1. **Serum:** Allow blood to clot at room temperature for 10–20 minutes, then centrifuge at 2000–3000 rpm for 20 minutes. 2. **Plasma:** Use EDTA or sodium citrate as anticoagulant, mix, and centrifuge similarly. 3. **Urine:** Centrifuge at 2000–3000 rpm for 20 minutes. 4. **Cell Culture Supernatant:** Centrifuge and collect the supernatant. For intracellular components, lyse cells via freezing-thawing and centrifuge again. 5. **Tissue Sample:** Homogenize in PBS, centrifuge, and collect the supernatant. 6. **Storage:** Avoid repeated freeze-thaw cycles. Store at -20°C if not tested immediately. Do not use samples containing NaN3. **Procedure Steps:** 1. Prepare standards in serial dilutions. 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 5 times with diluted washing buffer. 5. Add 50 µL of enzyme-labeled reagent to each well (except blank). 6. Incubate again for 30 minutes. 7. Add 50 µL of TMB A and B, incubate at 37°C for 15 minutes. 8. Stop the reaction with 50 µL of stop solution. 9. Measure OD at 450 nm within 15 minutes. **Notes:** - Equilibrate the kit to room temperature before use. - Avoid cross-contamination by using a new sealing film for each test. - Handle all reagents carefully and follow the manual strictly. - Discard all waste as biohazardous material. - Do not mix reagents from different batches. - Always prepare a standard curve and run duplicates for accuracy. **Performance:** - Linear correlation coefficient (R) ≥ 0.95 - Intra-batch CV < 9%, Inter-batch CV < 11% - Detection range: 0.2 IU/L – 6 IU/L **Technical Support:** We offer free technical guidance during working hours. Contact us for assistance with sample testing and optimization of results. **Note:** This kit is for research use only and not for diagnostic purposes.

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