**Monkey Alpha Interferon (IFN-α) ELISA Kit – Instructions for Use**
**Kit Specifications:**
This ELISA kit is available in 48-well or 96-well configurations. It includes:
- Standard dilution: 1.5 mL × 1 vial
- Enzyme standard reagent: 3 mL × 1 vial (48-well) / 6 mL × 1 vial (96-well)
- Sealing film: 2 pieces (48-well) / 2 pieces (96-well)
- Microplate: 1×48 or 1×96
- Sample diluent: 3 mL × 1 vial (48-well) / 6 mL × 1 vial (96-well)
- Developer A: 3 mL × 1 vial (48-well) / 6 mL × 1 vial (96-well)
- Chromogen B: 3 mL × 1 vial (48-well) / 6 mL × 1 vial (96-well)
- Stop solution: 3 mL × 1 vial (48-well) / 6 mL × 1 vial (96-well)
- Concentrated washing solution: 20 mL × 20 times or 20 mL × 30 times, depending on the configuration
**Storage Conditions and Expiration:**
- Storage: 2–8°C
- Shelf Life: 6 months from the date of receipt
**Purpose:**
This ELISA kit is designed to quantitatively determine the levels of Monkey Alpha Interferon (IFN-α) E in serum, plasma, urine, cell culture supernatants, tissue homogenates, and other biological fluids.
**Principle of the Assay:**
The kit uses a double-antibody sandwich ELISA method. The microtiter plate is pre-coated with a specific monoclonal antibody against IFN-α E. After adding the sample and standards, the target antigen binds to the immobilized antibody. An HRP-conjugated secondary antibody is then added, forming an immune complex. TMB substrate is used to develop the color, which is stopped by an acidic solution. The intensity of the color is directly proportional to the concentration of IFN-α E in the sample. Absorbance is measured at 450 nm using a microplate reader, and concentrations are calculated based on a standard curve.
**Sample Preparation and Requirements:**
1. **Serum:** Collect blood at room temperature, allow to clot for 10–20 minutes, centrifuge at 2000–3000 rpm for 20 minutes. Store supernatant at 2–8°C.
2. **Plasma:** Use EDTA or sodium citrate as anticoagulant. Mix well, centrifuge, and collect supernatant.
3. **Urine:** Collect in a sterile container, centrifuge, and collect supernatant.
4. **Cell Culture Supernatant:** Centrifuge after collection. For intracellular components, lyse cells via freezing/thawing, then centrifuge.
5. **Tissue Homogenate:** Weigh the tissue, add PBS, homogenize, centrifuge, and collect supernatant.
6. **Storage:** Avoid repeated freeze-thaw cycles. Store samples at -20°C if not tested immediately. Do not use samples containing NaN3.
**Operation Steps:**
1. **Standard Dilution:** Prepare a serial dilution of the standard (1800 ng/L to 150 ng/L).
2. **Sample Loading:** Add 40 µL of sample diluent and 10 µL of sample to each well (final dilution: 5x).
3. **Incubation:** Seal the plate and incubate at 37°C for 30 minutes.
4. **Washing:** Wash 5 times with diluted washing buffer.
5. **Enzyme Addition:** Add 50 µL of enzyme-labeled reagent to each well (except blank wells).
6. **Second Incubation:** Incubate again at 37°C for 30 minutes.
7. **Color Development:** Add 50 µL of TMB A and B, incubate at 37°C for 15 minutes.
8. **Stop Reaction:** Add 50 µL of stop solution to each well.
9. **Reading:** Measure OD450 within 15 minutes using a microplate reader.
**Notes:**
- Allow the kit to equilibrate to room temperature before use.
- Avoid cross-contamination; use a new sealing film for each test.
- Ensure accurate pipetting and maintain consistent timing during loading.
- Prepare a standard curve and run duplicates for better accuracy.
- If sample OD exceeds that of the highest standard, perform a preliminary dilution.
- All waste should be treated as biohazardous material.
- Do not mix reagents from different batches.
**Performance:**
- Linear regression correlation coefficient (R) ≥ 0.95
- Intra-assay CV < 9%, Inter-assay CV < 11%
- Detection range: 0.2 IU/L – 6 IU/L
**Service Commitment:**
We offer free technical support and guidance during working hours. Custom testing services are also available upon request.
**Delivery:**
Upon payment, the kit will be shipped promptly.
This kit is intended for research purposes only.
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