**Human Scd163 ELISA Kit – For the Quantitative In Vitro Determination of Human Soluble CD163 Concentrations in Serum, Plasma, Cerebrospinal Fluid, Tissue Homogenate, and Other Body Fluids**
**For Laboratory Research Use Only. NOT FOR USE IN DIAGNOSTIC PROCEDURES.**
Before beginning the assay, please read this entire package insert carefully to ensure proper use and accurate results.
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### **INTENDED USE AND TEST PRINCIPLE**
This Human Scd163 ELISA Kit is designed for research purposes only and is not intended for diagnostic or therapeutic use. The kit allows for the quantitative determination of soluble CD163 (Scd163) levels in various biological samples.
The test is based on a competitive enzyme-linked immunosorbent assay (ELISA) principle. A set of calibration standards is included with the kit, which are used to generate a standard curve. By comparing the optical density (OD) of the sample to this curve, the concentration of Scd163 in the sample can be accurately determined.
A stop solution is added at the end of the procedure, causing the color to change from blue to yellow. The intensity of the color is directly proportional to the amount of Scd163 present in the sample.
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### **SAMPLE COLLECTION AND STORAGE**
- **Serum:** Collect using a serum separator tube. Allow samples to clot for 2 hours at room temperature or overnight at 4°C before centrifugation at 2000×g for 20 minutes. Remove serum and analyze immediately or aliquot and store at -20°C. Avoid repeated freeze-thaw cycles.
- **Plasma:** Collect using heparin as an anticoagulant. Centrifuge within 30 minutes at 2000×g, at 2–8°C. Avoid repeated freeze-thaw cycles.
- **Cell Culture Supernatants, Tissue Homogenates, and Other Biological Fluids:** Centrifuge to remove particulates and analyze immediately or store at -20°C. Avoid repeated freeze-thaw cycles.
**Note:** Ensure proper centrifugation and avoid hemolysis or particle contamination in all samples.
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### **MATERIALS REQUIRED BUT NOT SUPPLIED**
1. Incubator set at 37°C
2. Microplate reader capable of measuring absorbance at 450 nm
3. Precision pipettes, disposable tips, and absorbent paper
4. Distilled or deionized water
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### **REAGENTS PROVIDED (Stored at 2–8°C)**
| Reagent Name | 96 Determinations | 48 Determinations |
|--------------|-------------------|-------------------|
| MicroElisa Stripplate | 12 x 8 strips | 12 x 4 strips |
| Standard (6 vials) | 0.5 ml/vial | 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 |
**Note:**
- Standard concentrations: 160, 80, 40, 20, 10, 5 ng/mL.
- If sample values exceed the highest standard, dilute with Sample Diluent and repeat the assay.
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### **PRECAUTIONS**
1. Do not mix reagents from different kit lots. All components are calibrated for optimal performance.
2. Allow all reagents and samples to reach room temperature (20–25°C) before use. Do not use water baths to thaw.
3. Do not use reagents past their expiration date.
4. Always use fresh pipette tips for each step.
5. Handle all plasma samples as potentially hazardous. Wear gloves during the procedure.
6. Dispose of all waste properly after inactivation (allow 30 minutes).
7. Substrate B contains 20% acetone; keep away from heat or flame.
8. Store unused strips in sealed bags with desiccant at 2–8°C.
9. Do not remove microtiter plates from storage bags until needed.
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### **REAGENT PREPARATION AND STORAGE**
**Wash Solution (1X):** Dilute 1 volume of 20X Wash Solution with 19 volumes of distilled or deionized water. Store at 2–8°C for up to 1 month.
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### **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 receives no addition.
3. Add 100 µL of HRP-conjugate reagent to all wells except blank. Cover with adhesive strip and incubate for 60 minutes at 37°C.
4. Wash plate 4 times manually or via automated washer.
- Manual: Fill each well with 1X Wash Solution, aspirate, and repeat 4 times.
- Automated: Aspirate and wash with 350 µL/well.
5. After washing, 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.
6. Add 50 µL of Stop Solution to each well. Color changes from blue to yellow. If uneven, gently tap to mix.
7. Measure OD at 450 nm. Plot standard curve using average OD values. Subtract blank OD from all readings.
8. Determine sample concentration by matching OD to the standard curve.
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### **PERFORMANCE CHARACTERISTICS**
- **Intra-assay CV (%) and Inter-assay CV (%) < 15%**
- **Assay Range:** 5 ng/mL – 160 ng/mL
- **Sensitivity:** <1.0 ng/mL
- **No cross-reactivity or interference observed**
- **Storage:** 2–8°C (frequent use); 6 months at -20°C
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### **NOTES**
- Ensure consistent pipetting and incubation conditions for reliable results.
- Each user should prepare their own standard curve.
- Follow all safety guidelines when handling biological samples.
- This kit is for research use only and should not be used in clinical settings.
**Please read all instructions carefully before use.**
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