**Goat IgG ELISA Kit – For the Quantitative In Vitro Determination of Goat Immunoglobulin G in Serum, Plasma, Cerebrospinal Fluid, Tissue Homogenate, and Other Body Fluids**
*For Laboratory Research Use Only. Not Intended for Diagnostic Procedures.*
Before using this product, please read the entire package insert carefully. This ELISA kit is designed for research purposes only and should not be used in clinical or diagnostic settings. The test is based on the enzyme-linked immunosorbent assay (ELISA) principle, which allows for the accurate quantification of Goat IgG levels in various biological samples.
The kit includes a set of calibration standards that are run alongside the samples to generate a standard curve. By measuring the optical density (OD) at 450 nm, the concentration of IgG in each sample can be determined by comparing its OD value to the standard curve. A stop solution is provided to halt the reaction, changing the color from blue to yellow, making it easier to measure results.
**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 centrifuging at 2000×g for 20 minutes. Assay immediately or store at -20°C in aliquots. Avoid repeated freeze-thaw cycles.
- **Plasma**: Collect 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.
- **Cell Culture Supernatants, Tissue Homogenates, and Other Biological Fluids**: Centrifuge to remove particulates. Assay immediately or store at -20°C. Ensure no hemolysis or contamination occurs.
**Materials Required but Not Supplied**
- 37°C incubator
- Microplate reader capable of measuring absorbance at 450 nm
- 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.
| Reagent | 96 Determinations | 48 Determinations |
|---------|-------------------|-------------------|
| MicroELISA Strip Plate | 12×8 strips | 12×4 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: 480, 240, 120, 60, 30, 15 μg/mL
- If sample values exceed the highest standard, dilute with Sample Diluent and repeat the assay.
**Precautions**
1. Use only the reagents provided. Do not mix reagents from different kits.
2. Allow all reagents and samples to reach room temperature (20–25°C) before use.
3. Do not use reagents past their expiration date.
4. Use only deionized or distilled water for dilutions.
5. Keep unused microplate strips in sealed bags with desiccant at 2–8°C.
6. Use fresh pipette tips for each transfer to avoid cross-contamination.
7. Handle all biological materials with care, assuming potential infectious risk.
8. Dispose of all waste following proper inactivation protocols.
9. Liquid waste: Add sodium hypochlorite to a final concentration of 1.0% and let stand for 30 minutes before disposal.
10. Substrate solutions may be easily contaminated; discard if discolored.
11. Chromogen B contains 20% acetone—keep away from heat and flame.
**Reagent Preparation**
- **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. Run standards and samples in duplicate.
2. Add 50 μl of standard or sample to each well. Leave blank well empty.
3. Add 100 μl of HRP-conjugate reagent to all wells except the blank. Cover with adhesive strip and incubate at 37°C for 60 minutes.
4. Wash the plate 4 times manually or automatically.
- **Manual Washing**: Aspirate contents, fill with 1X Wash Solution, and repeat four times. Blot dry after final wash.
- **Automated Washing**: Follow manufacturer instructions.
5. Add 50 μl of Chromogen A and 50 μl of Chromogen B to each well. Incubate at 37°C for 15 minutes, protecting from light.
6. Add 50 μl of Stop Solution to each well. Read OD at 450 nm within 15 minutes.
**Calculation**
- Plot average OD values (450 nm) against standard concentrations.
- Subtract blank OD from all readings.
- Use graph paper or software to construct the standard curve.
- Locate the sample OD on the Y-axis and draw a line to intersect the curve. Read the corresponding IgG concentration on the X-axis.
- Each user should generate their own standard curve due to possible variations in technique.
**Sensitivity & Specificity**
- Sensitivity: <1.0 μg/mL
- Cross-reactivity: No significant cross-reactivity with other immunoglobulins.
**Storage**
- 2–8°C for frequent use; 6 months at -20°C.
Always follow good laboratory practices and handle all reagents with care. This kit is intended solely for research purposes and must not be used in clinical diagnostics.
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