Human TSP-1 ELISA Kit

**Human TSP-1 ELISA Kit – For the quantitative in vitro determination of Human thrombospondin 1 concentrations in serum, plasma, celiac fluid, tissue homogenate, and body fluids. FOR LABORATORY RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES.** This package insert must be read thoroughly before using the ELISA kit. The kit is designed for research purposes only and should not be used in diagnostic or therapeutic procedures. The Stop Solution changes the color from blue to yellow, and the intensity of the color is measured at 450 nm using a spectrophotometer. To determine the concentration of TSP-1 in the sample, this TSP-1 ELISA Kit includes a set of calibration standards. These standards are assayed alongside the samples to generate a standard curve of optical density (OD) versus TSP-1 concentration. The TSP-1 levels in the samples are then calculated 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 before centrifugation for 20 minutes at approximately 2000×g. Remove the serum and assay immediately, or aliquot and store at -20°C. Avoid repeated freeze-thaw cycles. - **Plasma:** Collect plasma using heparin as an anticoagulant. Centrifuge within 30 minutes of collection at 2000×g for 30 minutes at 2–8°C. Store at -20°C. Avoid repeated freeze-thaw cycles. - **Cell culture supernatants, tissue homogenates, and other biological fluids:** Remove particulates by centrifugation and assay immediately, or aliquot and store at -20°C. Ensure adequate centrifugation and avoid hemolysis or granules. **Materials Required but Not Supplied:** - 37°C incubator - Standard microplate reader capable of measuring absorbance at 450 nm - Precision pipettes, disposable pipette tips, and absorbent paper - Distilled or deionized water **Reagents Provided (Stored at 2–8°C):** - MicroELISA strip plate (12x8 strips / 12x4 strips) - Standard (6 vials, 0.5 ml/vial) - Sample diluent (6.0 ml / 3.0 ml) - HRP-Conjugate reagent (10.0 ml / 5.0 ml) - 20X Wash solution (25 ml / 15 ml) - Chromogen Solution A (6.0 ml / 3.0 ml) - Chromogen Solution B (6.0 ml / 3.0 ml) - Stop Solution (6.0 ml / 3.0 ml) - Closure plate membrane (2 / 2) - User Manual (1 / 1) - Sealed bags (1 / 1) **Notes:** - Standard concentrations: 2000, 1000, 500, 250, 125, 62.5 ng/mL - If sample values exceed the highest standard, dilute with Sample Diluent and repeat the test. - Do not use the kit beyond its expiration date. - All reagents should reach room temperature (20–25°C) before use. - Do not thaw samples or reagents in water baths. - Only use deionized or distilled water for dilution. - Keep unused strips in the sealed bag with desiccant at 2–8°C. - Use fresh pipette tips for each transfer to avoid contamination. - Treat all disposable materials as potentially hazardous. - Follow proper disposal procedures for all samples. - Allow liquid to stand for at least 30 minutes to inactivate viruses before disposal. - Substrate Solution may be easily contaminated. Avoid exposure to heat or flame. - Ensure all reagents are at room temperature before starting the assay. **Reagent Preparation and Storage:** - **Wash Solution (1X):** Dilute 1 volume of 20X Wash Solution with 19 volumes of deionized or distilled water. Store at 2–8°C for up to one month. **Assay Procedure:** 1. Prepare all reagents before starting the assay. 2. Add 50 µL of standard or sample to appropriate wells (excluding blank well), cover with adhesive strip, and incubate for 60 minutes at 37°C. 3. Wash the microtiter plate 4 times manually or using automated washing. 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 using a microplate reader. **Data Interpretation:** - Plot average OD values (450 nm) of the six standards on the X-axis. - Subtract blank OD from all readings before interpretation. - Construct a standard curve using graph paper or software. - Locate the OD value on the Y-axis, draw a horizontal line to the curve, and then a vertical line to the X-axis to find the corresponding TSP-1 concentration. **Performance Characteristics:** - Intra-assay CV (%) and Inter-assay CV (%) < 15% - Assay range: 62.5 ng/mL – 2000 ng/mL - Sensitivity: < 10 ng/mL - Cross-reactivity: No significant cross-reactivity observed - Storage: 2–8°C (for frequent use); -20°C for long-term storage (up to 6 months) **Important Safety Information:** - Handle all samples and reagents with care. - Follow good laboratory practices to minimize risk of contamination or exposure. - Always wear appropriate personal protective equipment (PPE). **Note:** This kit is intended for research use only and should not be used in clinical diagnostics.

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