Human S-100B ELISA Kit
2025-10-11 09:08:49
**Human S-100B ELISA Kit – For the Quantitative In Vitro Determination of Human Soluble Protein-100B Concentrations in Serum, Plasma, Cerebrospinal Fluid, Tissue Homogenate, and Body Fluids**
**For Laboratory Research Use Only. Not for Use in Diagnostic Procedures.**
This package insert must be read carefully before use. The Human S-100B ELISA Kit is intended for laboratory research purposes only and should not be used for diagnostic or therapeutic applications. The kit is based on the Enzyme-Linked Immunosorbent Assay (ELISA) principle, which allows for the precise quantification of S-100B protein levels in biological samples.
The reaction is terminated by adding a Stop Solution, which changes the color from blue to yellow. The intensity of the yellow color is measured at 450 nm using a spectrophotometer. A standard curve is generated using a set of calibration standards provided in the kit. These standards are run alongside the samples, allowing the operator to determine the concentration of S-100B in unknown samples by comparing their optical density (OD) values to the standard curve.
**Sample Collection and Storage**
- **Serum**: Collect blood in a serum separator tube. Allow clotting 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.
- **Cell Culture Supernatants, Tissue Homogenates, and Other Biological Fluids**: Remove particulates by centrifugation. Analyze immediately or aliquot and store at -20°C. Ensure no hemolysis or granules are present in the sample.
**Materials Required but 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**
All reagents are stored at 2–8°C. Check the expiration date on the label.
- Microtiter Strip Plate: 12×8 strips 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 units
- User Manual: 1 unit
- Sealed Bags: 1 unit
**Notes**
- Standard concentrations: 640, 320, 160, 80, 40, 20 pg/mL.
- If sample values exceed the highest standard, dilute with Sample Diluent.
- Do not use any reagents beyond their expiration date.
- Use only deionized or distilled water for dilutions.
- Keep microtiter plates in sealed bags until use. Unused strips should be stored at 2–8°C with desiccant.
- Use fresh pipette tips for each transfer to prevent cross-contamination.
**Safety and Disposal**
- Disposable gloves must be treated as potentially hazardous and disposed of accordingly.
- All samples and reagents should be considered potentially infectious. Follow good laboratory practices.
- Liquid waste should be mixed with sodium hypochlorite to a final concentration of 1.0% and allowed to stand for at least 30 minutes before disposal.
- Substrate Solution B contains 20% acetone; keep away from heat and flame.
**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 1 month.
**Assay Procedure**
1. Prepare all reagents before starting. Run standards and samples in duplicate.
2. Add 50 µl of standard or sample to appropriate wells. Blank well: no addition.
3. Add 100 µl of HRP-conjugate reagent to all wells except blank. Cover with adhesive strip. Incubate for 60 minutes at 37°C.
4. Wash the plate 4 times manually or using an automated washer.
- Manual: Fill each well with 1X Wash Solution, aspirate, repeat 4 times. Dry by blotting.
- Automated: Aspirate and wash four times with 1X Wash Buffer. Adjust brush and fill volume to 350 µL/well.
5. Add 50 µl of Chromogen A and 50 µl of Chromogen B to each well. Mix gently and incubate for 15 minutes at 37°C, protecting from light.
6. Add 50 µl of Stop Solution to each well. Color changes from blue to yellow. Gently tap if uneven.
7. Read OD at 450 nm. Generate a standard curve by plotting average OD values against known concentrations.
**Results Interpretation**
- Calculate mean OD for each standard and sample. Subtract blank OD before interpretation.
- Plot standard curve and determine sample concentration by interpolation.
- Intra-assay and inter-assay CV% < 15%.
- Assay range: 20 pg/mL – 640 pg/mL.
- Sensitivity: <20 pg/mL.
- Cross-reactivity: No significant cross-reactivity observed.
**Storage**
- Store at 2–8°C for frequent use. Long-term storage: -20°C. Shelf life: 6 months.
**Important Notes**
- Always allow reagents to reach room temperature (20–25°C) before use.
- Avoid using water baths for thawing.
- Follow all safety guidelines and dispose of waste properly.
This kit is designed for accurate and reliable detection of S-100B in various biological matrices. It is essential to follow all instructions carefully to ensure consistent and reproducible results.
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