Human Apoptosis Related Factor Ligand (FASL) ELISA Kit Instructions

**Human Apoptosis-Related Factor Ligand (FASL) ELISA Kit** **For the quantitative in vitro determination of Human FASL concentrations in serum, plasma, cerebrospinal fluid, tissue homogenate, and other body fluids.** **Intended Use:** This FASL ELISA Kit is designed for laboratory research use only and is not intended for diagnostic or therapeutic purposes. The kit enables the accurate measurement of FASL levels using a colorimetric enzyme-linked immunosorbent assay (ELISA). **Test Principle:** The FASL ELISA Kit utilizes a competitive binding immunoassay. A standard curve is generated by measuring optical density (OD) at 450 nm using a microplate reader. The concentration of FASL in unknown samples is determined by comparing their OD values to the standard curve. **Sample Collection and Storage:** - **Serum:** Use a serum separator tube. Allow samples to clot for 2 hours at room temperature or overnight at 4°C. Centrifuge at 2000×g for 20 minutes. Store at -20°C. Avoid repeated freeze-thaw cycles. - **Plasma:** Collect using heparin as an anticoagulant. Centrifuge within 30 minutes at 2000×g for 30 minutes at 2–8°C. Store at -20°C. - **Cell culture supernatants, tissue homogenates, and other biological fluids:** Centrifuge to remove particulates. Assay immediately or store at -20°C. Avoid freeze-thaw cycles. **Materials Required (Not Supplied):** - 37°C incubator - Microplate reader capable of measuring absorbance at 450 nm - Precision pipettes, disposable tips, and absorbent paper - Distilled or deionized water **Reagents Provided:** - Microtiter Strip Plate (12×8 or 12×4 strips) - Standard (6 vials, 0.5 ml/vial) – 2000, 1000, 500, 250, 125, 62.5 pg/ml - Sample Diluent (6.0 ml or 3.0 ml) - HRP-Conjugate Reagent (10.0 ml or 5.0 ml) - 20X Wash Solution (25 ml or 15 ml) - Chromogen Solutions A & B (6.0 ml or 3.0 ml each) - Stop Solution (6.0 ml or 3.0 ml) - Closure Plate Membrane (2 pieces) - User Manual and Sealed Bags **Precautions:** - Do not mix reagents from different kit lots. - Allow all reagents to reach room temperature before use. - Never use reagents beyond their expiration date. - Always use deionized or distilled water for dilutions. - Handle all biological samples with care, assuming potential infectious risk. - Dispose of waste properly after inactivation. **Reagent Preparation:** Dilute 1 volume of 20X Wash Solution with 19 volumes of deionized water. Store at 2–8°C for up to one month. **Assay Procedure:** 1. Prepare all reagents. Add 50 µl of standards or samples in duplicate to the microtiter plate. 2. Add 100 µl of HRP-conjugate reagent to each well (except blank), cover, and incubate for 60 minutes at 37°C. 3. Wash the plate 4 times with 1X Wash Solution. 4. Add 50 µl of Chromogen A and 50 µl of Chromogen B to each well. Incubate for 15 minutes at 37°C, protected from light. 5. Add 50 µl of Stop Solution to each well. Read OD at 450 nm within 15 minutes. **Calculation of Results:** - Plot average OD values against standard concentrations to generate a standard curve. - Subtract blank OD from all measurements. - Determine sample concentration by interpolating OD values on the standard curve. - Intra-assay and inter-assay CV% < 15%. - Sensitivity: <10 pg/ml. - Cross-reactivity: No significant cross-reaction with other ligands. **Storage:** - Store at 2–8°C for frequent use. - For long-term storage, keep at -20°C. - Shelf life: 6 months when stored properly. **Note:** This product is for research use only. Follow all safety guidelines and handle all reagents with care. Read the entire manual before use.

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