**Sheep IL-1 (IL-1) ELISA Kit – Instructions for Use**
**Kit Specifications:**
This Sheep IL-1 ELISA Kit is available in 48-well or 96-well configurations. It is designed for the quantitative determination of Interleukin-1 (IL-1) in biological samples such as serum, plasma, urine, cell culture supernatants, and tissue homogenates. The kit includes all necessary reagents for a complete assay.
**Standard Dilution:**
- 1.5 mL × 1 vial (for standard preparation)
- Enzyme Standard Reagent: 3 mL × 1 vial (48-well) / 6 mL × 1 vial (96-well)
**Storage Conditions & Shelf Life:**
- Store at 2–8°C
- Shelf life: 6 months from the date of manufacture
**Kit Composition:**
- Sealing Film: 2 pieces (48-well) / 2 pieces (96-well)
- Standard: 0.5 mL × 1 vial (2700 ng/L)
- Enzyme Standard: 1×48 / 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)
- Wash Solution: 3 mL × 1 vial (48-well) / 6 mL × 1 vial (96-well)
- Concentrated Wash Solution: 20 mL × 20 times / 20 mL × 30 times (per bottle)
**Principle of Operation:**
The kit utilizes the sandwich ELISA method. Microtiter plates are pre-coated with anti-IL-1 antibodies. After incubation with the sample, IL-1 binds to the immobilized antibody. HRP-conjugated secondary antibody then binds to the captured IL-1, forming an immune complex. TMB substrate is added, producing a blue color that turns yellow upon acid termination. The intensity of the color is directly proportional to the IL-1 concentration in the sample. Absorbance is measured at 450 nm, and concentrations are calculated using a standard curve.
**Purpose:**
This ELISA kit is intended for research use only. It is used to quantify IL-1 levels in various biological fluids, including serum, plasma, urine, and cell culture supernatants.
**Sample Preparation Guidelines:**
- **Serum:** Allow blood to clot at room temperature for 10–20 minutes, then centrifuge at 2000–3000 rpm for 20 minutes. Collect the supernatant.
- **Plasma:** Use EDTA or sodium citrate as anticoagulant. Mix well and centrifuge as above.
- **Urine:** Centrifuge at 2000–3000 rpm for 20 minutes. Collect the supernatant.
- **Cell Culture Supernatant:** Centrifuge to remove debris. For intracellular components, lyse cells by freezing/thawing and centrifuge again.
- **Tissue Homogenate:** Weigh the tissue, add PBS, homogenize, and centrifuge. Collect the supernatant.
**Important Notes:**
- Avoid repeated freeze-thaw cycles.
- Do not use samples containing NaN3, as it inhibits HRP activity.
- Always prepare a standard curve with duplicate wells.
- Keep all reagents away from light.
- Follow instructions strictly. Results must be read on a microplate reader.
- Discard all waste as biohazardous material.
**Performance Characteristics:**
- Correlation coefficient (R²) ≥ 0.95
- Intra-batch CV < 9%, Inter-batch CV < 11%
- Detection range: 0.2 IU/L – 6 IU/L
**Service Commitment:**
We provide free technical support during working hours. Custom testing services are available upon request to ensure accurate results.
**Usage Steps Summary:**
1. Prepare standards and samples.
2. Add to microplate and incubate.
3. Wash plate thoroughly.
4. Add enzyme conjugate and incubate.
5. Add TMB substrate and develop color.
6. Stop reaction and measure OD at 450 nm.
7. Calculate concentration using standard curve.
**Safety & Handling:**
All reagents should be handled with care. Wear gloves and lab coat. Avoid contact with skin and eyes.
**Note:** This kit is for research purposes only and not for diagnostic use.
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