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Human DHEA-S ELISA Kit
**Human DHEA-S ELISA Kit – For the Quantitative In Vitro Determination of Human Dehydroepiandrosterone Sulfate (DHEA-S) in Serum, Plasma, Cerebrospinal Fluid, Tissue Homogenate, and Other Biological Fluids**
**For Laboratory Research Use Only. Not for Diagnostic Procedures.**
This ELISA kit is designed for the accurate quantification of DHEA-S levels in various biological samples. The Stop Solution changes the color from blue to yellow, and the optical density (OD) is measured at 450 nm using a spectrophotometer. A set of calibration standards is included to generate a standard curve, allowing for precise determination of DHEA-S concentrations in unknown samples.
**Sample Collection and Storage:**
- **Serum:** Use serum separator tubes. Allow samples to clot for 2 hours at room temperature or overnight at 4°C before centrifuging at 2000×g for 20 minutes. Remove serum and assay immediately or store at -20°C. Avoid repeated freeze-thaw cycles.
- **Plasma:** Collect using heparin as an anticoagulant. Centrifuge within 30 minutes at 2000×g (2–8°C). Avoid freeze-thaw cycles.
- **Cell Culture Supernatants, Tissue Homogenates, and Other Fluids:** Centrifuge to remove particulates and assay immediately or store at -20°C. Ensure no hemolysis or granules are present.
**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 (Stored at 2–8°C):**
- Microtiter Strip Plate (12×8 or 12×4 strips)
- Standards (6 vials, 0.5 ml/vial)
- 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 Solution A (6.0 ml or 3.0 ml)
- Chromogen Solution B (6.0 ml or 3.0 ml)
- Stop Solution (6.0 ml or 3.0 ml)
- Closure Plate Membrane (2 pieces)
- User Manual and Sealed Bags
**Standard Concentrations:** 800, 400, 200, 100, 50, 25 pg/ml
**Note:** If sample values exceed the highest standard, dilute with Sample Diluent and repeat the assay.
**Precautions:**
- Do not mix reagents from different kits.
- Allow all reagents to reach room temperature (20–25°C) before use.
- Do not use water baths to thaw samples or reagents.
- Do not use beyond the expiration date.
- Use only deionized or distilled water.
- Keep unused strips in sealed bags with desiccant.
- Wear gloves when handling biological samples.
- Dispose of waste properly after inactivation for 30 minutes.
- Avoid contamination of substrate solutions.
- Chromogen B contains 20% acetone; keep away from heat or flame.
**Reagent Preparation:**
- Wash Solution (1X): Dilute 1 volume of 20X solution with 19 volumes of water. Store at 2–8°C for 1 month.
**Assay Procedure:**
1. Prepare all standards and samples in duplicate.
2. Add 50 μl of standard or sample to wells. Blank well receives no addition.
3. Add 100 μl of HRP-conjugate reagent to all wells except blank. Incubate for 60 minutes at 37°C.
4. Wash plate 4 times manually or automatically.
5. Add 50 μl each of Chromogen A and B. Incubate for 15 minutes at 37°C, protected from light.
6. Add 50 μl Stop Solution. Color changes from blue to yellow. Gently mix if uneven.
7. Measure OD at 450 nm. Calculate mean OD for each standard and sample. Subtract blank value before plotting.
**Standard Curve:**
Plot average OD vs. DHEA-S concentration. Locate sample OD on Y-axis and read corresponding concentration from the curve.
**Performance Characteristics:**
- Intra-assay CV <15%, Inter-assay CV <15%
- Assay range: 25 pg/ml – 800 pg/ml
- Sensitivity: <10 pg/ml
- Cross-reactivity: No significant interference
- Storage: 2–8°C (frequent use); 6 months at -20°C
**Important Note:** This kit is intended for research use only. It should not be used in diagnostic procedures. Always follow good laboratory practices and handle all samples with care.