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Rat eosinophil chemotactic factor (ECF) elisa kit instruction manual
**Rat Eosinophil Chemotactic Factor (ECF) ELISA Kit – User Manual**
**Kit Specifications:**
This ELISA kit is available in 48-well or 96-well configurations. The standard reagent is provided as 1.5 mL × 1 vial, with the enzyme standard solution available in 3 mL × 1 vial (for 48-well) or 6 mL × 1 vial (for 96-well). The kit is intended for research use only.
**Principle of Operation:**
The kit employs a sandwich ELISA method to quantify rat ECF levels in biological samples. A microplate pre-coated with purified anti-ECF antibodies is used. After incubation with the sample and HRP-labeled anti-ECF antibody, the complex is developed using TMB substrate. The reaction is stopped with an acidic solution, and the color intensity is measured at 450 nm. The concentration of ECF is determined by comparing the OD values to a standard curve.
**Kit Components:**
- Sealing film: 2 pieces (48-well) / 2 pieces (96-well)
- Standard: 0.5 mL × 1 vial (2700 ng/L)
- Enzyme standard: 3 mL × 1 vial (48-well) / 6 mL × 1 vial (96-well)
- Sample diluent: 3 mL × 1 vial / 6 mL × 1 vial
- TMB substrate A: 3 mL × 1 vial
- TMB substrate B: 3 mL × 1 vial
- Washing buffer: 20 mL × 20 times / 20 mL × 30 times
- Stop solution: 3 mL × 1 vial
**Storage Conditions:**
Store all components at 2–8°C. The kit is valid for 6 months from the date of receipt.
**Sample Preparation:**
- **Serum:** Allow blood to clot at room temperature, then centrifuge at 2000–3000 rpm for 10–20 minutes.
- **Plasma:** Use EDTA or sodium citrate as anticoagulant. Centrifuge after mixing.
- **Urine:** Centrifuge at 2000–3000 rpm for 20 minutes.
- **Cell culture supernatant:** Centrifuge after cell lysis.
- **Tissue homogenate:** Homogenize in PBS, centrifuge, and collect the supernatant.
**Procedure Summary:**
1. Prepare standard dilutions in a serial manner.
2. Add sample and standards to the microplate.
3. Incubate at 37°C for 30 minutes.
4. Wash the plate 5 times with diluted washing buffer.
5. Add HRP-conjugated antibody and incubate again.
6. Add TMB substrate and develop for 15 minutes.
7. Stop the reaction with stop solution.
8. Measure OD at 450 nm using a microplate reader.
9. Calculate concentrations using a standard curve.
**Notes:**
- Ensure all reagents are equilibrated to room temperature before use.
- Avoid repeated freezing and thawing of samples.
- Do not mix reagents from different batches.
- Keep TMB away from light.
- All waste materials should be treated as biohazardous.
**Performance Characteristics:**
- Linear regression correlation coefficient (R) ≥ 0.95
- Intra-assay CV < 9%, Inter-assay CV < 11%
- Detection range: 0.2 IU/L – 6 IU/L
**Technical Support:**
Free assistance is available during working hours. We also offer free sample testing to ensure optimal results. Please contact us for more details.
**Delivery & Warranty:**
The kit will be delivered within the agreed time after payment. For any questions, feel free to reach out.
This manual provides detailed guidance on using the Rat Eosinophil Chemotactic Factor (ECF) ELISA Kit. Following these instructions ensures accurate and reliable results in your research.