Key Features
- Exceptional Specificity for Equine IL-8: No cross-reactivity with human, mouse, rat, bovine, or canine IL-8. No interference from other equine soluble structural analogues.
- High Sensitivity & Defined Range: Minimum detectable concentration <1.0 pg/mL; quantitative range 1.0–240 pg/mL, supported by a six-point standard curve (0, 15, 30, 60, 120, 240 pg/mL).
- Recombinant Protein Standards: Produced in a eukaryotic expression system, ensuring native-like conformation and optimal immunoreactivity.
- Validated Sample Types: Equine serum, EDTA/citrate/heparin plasma, cell culture supernatant, and tissue homogenates.
- Comprehensive Tissue Processing Protocol: Includes mandatory protease inhibitor use, endogenous peroxidase inactivation for high-background tissues (liver, kidney, etc.), and low-temperature centrifugation to preserve analyte integrity.
- Built-in 5× Sample Dilution: The assay automatically dilutes samples 5-fold in the well (10 µL sample + 40 µL Sample Diluent). Multiply the interpolated result by 5 to obtain the actual concentration.
- One-Step Sandwich ELISA: Simultaneous incubation of sample/standard and HRP-detection antibody; total assay time approximately 2 hours.
- Reliable Performance: Intra- and inter-assay CV <15%; standard curve linearity R ≥0.9900.
- Flexible Formats: 48-well and 96-well configurations with removable strips; up to 88 samples per plate (single wells).
- Storage & Stability: 6-month shelf life at 2–8°C, protected from light and moisture; cold-chain shipping.
Product Description
Interleukin-8 (IL-8, CXCL8) is a potent neutrophil chemoattractant and a key mediator of acute inflammation. In horses, IL-8 is critically involved in the pathogenesis of inflammatory airway disease, colic, endometritis, laminitis, and systemic inflammatory response syndrome. Accurate measurement of equine IL-8 is essential for research into innate immunity, infectious disease, and tissue injury in the horse.
The Yanda-HORSE-IL-8 ELISA Kit is a one-step double-antibody sandwich ELISA. A microplate pre-coated with an antibody specific for equine IL-8 captures the target from samples and standards. An HRP-conjugated detection antibody is added simultaneously in a single incubation step. After washing, TMB substrate produces a colorimetric signal proportional to the IL-8 concentration. Absorbance is measured at 450 nm, and results are calculated from the standard curve.
Sample Collection & Preparation
General Guidelines for Liquid Samples
| Sample Type | Procedure |
|---|---|
| Serum | Collect blood in pyrogen/endotoxin-free tubes. Let clot naturally, then centrifuge at 3000 rpm for 10 min. Separate serum immediately. |
| Plasma | Use EDTA, citrate, or heparin as anticoagulant. Centrifuge at 3000 rpm for 30 min; collect supernatant. |
| Cell Culture Supernatant | Centrifuge at 3000 rpm for 10 min to remove cells and debris. |
Tissue Homogenate Protocol (Critical for Reliable Results)
Mandatory Use of Protease Inhibitors: Tissues contain abundant endogenous proteases that rapidly degrade IL-8. Freshly add a protease inhibitor cocktail (or 1 mM PMSF) to the ice-cold homogenization buffer before use.
Endogenous Peroxidase Inactivation: Tissues with high endogenous peroxidase activity (e.g., liver, kidney, pancreas) can produce false-positive signals. After sample preparation, treat the supernatant with 1% H₂O₂ for 15 minutes at room temperature to inactivate these peroxidases before the assay.
Standardized Homogenization Steps:
- Rinse: Quickly rinse fresh tissue with ice-cold PBS to remove residual blood.
- Homogenize: Add ice-cold lysis buffer or PBS (containing fresh protease inhibitors) at a ratio of 100 mg tissue : 1 mL buffer. Homogenize thoroughly on ice.
- Incubate on Ice: Let the homogenate sit on ice for 30 minutes, mixing occasionally.
- Centrifuge: Centrifuge at 12,000 × g for 5–10 minutes at 4°C. Carefully collect the clear middle supernatant for testing.
Storage for All Samples:
- If not assayed immediately, aliquot in single-use volumes and store at -20°C. Avoid repeated freeze-thaw cycles. Avoid sample hemolysis.Thaw completely at room temperature and mix gently before use. Do not use samples containing sodium azide (NaN₃), as it inhibits HRP activity.
Critical Dilution Factor
The assay protocol applies a 5-fold dilution directly in the well (10 µL sample + 40 µL Sample Diluent). The concentration interpolated from the standard curve represents the diluted sample. Multiply the obtained value by 5 to determine the actual IL-8 concentration in the original specimen.
The S0 standard (0 pg/mL) serves as the blank/negative control.
Note on Normal Serum Levels: Circulating IL-8 concentrations in healthy horses can be very low and may approach the lower detection limit of the assay. For baseline studies, we recommend performing pilot experiments to establish the detection frequency and expected range in your specific population and sample matrix.
Technical Specifications
| Parameter | Specification |
|---|---|
| Standard Concentrations (S0–S5) | 0, 15, 30, 60, 120, 240 pg/mL |
| Quantitative Range | 1.0 – 240 pg/mL |
| Analytical Sensitivity | < 1.0 pg/mL |
| Accuracy (Linearity) | Standard curve R ≥ 0.9900 |
| Precision | Intra-assay CV < 15%; Inter-assay CV < 15% |
| Specificity | Highly specific for equine IL-8. No cross-reactivity with human, mouse, rat, bovine, or canine IL-8. No interference from other soluble structural analogues. |
| Sample Dilution Factor | 5× (automatically performed in the well) |
| Shelf Life | 6 months |
| Storage & Transport | 2–8°C, protected from light and moisture; cold-chain shipping |
Representative Standard Curve
For spike-recovery, dilution linearity, and other validation data, please refer to the product instruction manual or contact technical support.
Kit Components
96-Well Format
| Component | Quantity | Notes |
|---|---|---|
| Microelisa Stripplate | 12 strips × 8 wells | Pre-coated with anti-equine IL-8 antibody |
| Standard (0.3 mL/vial) | 6 vials | Ready-to-use liquid, 0–240 pg/mL |
| Sample Diluent | 6 mL | – |
| HRP-Conjugated Detection Antibody | 10 mL | Ready-to-use |
| 20× Wash Buffer | 25 mL | Dilute to 1× before use |
| Substrate A | 6 mL | – |
| Substrate B | 6 mL | – |
| Stop Solution | 6 mL | – |
| Plate Sealers | 2 sheets | – |
| Instruction Manual | 1 copy | – |
| Self-sealing Bag | 1 piece | For storing unused strips |
48-Well Format
| Component | Quantity | Notes |
|---|---|---|
| Microelisa Stripplate | 12 strips × 4 wells | Pre-coated, removable |
| Standard (0.3 mL/vial) | 6 vials | Ready-to-use liquid |
| Sample Diluent | 3 mL | – |
| HRP-Conjugated Detection Antibody | 5 mL | Ready-to-use |
| 20× Wash Buffer | 15 mL | Dilute to 1× |
| Substrate A | 3 mL | – |
| Substrate B | 3 mL | – |
| Stop Solution | 3 mL | – |
| Plate Sealers | 2 sheets | – |
| Instruction Manual | 1 copy | – |
| Self-sealing Bag | 1 piece | – |
Note: The 20× Wash Buffer may crystallize during cold storage. Warm in a water bath until crystals dissolve completely before diluting to 1×.
Assay Procedure Summary
- Equilibrate: Allow the sealed foil pouch to reach room temperature for 20 minutes. Remove required strips; reseal unused strips and store at 4°C.
- Add Standards & Samples:
- Standard wells: 50 µL of each standard.
- Sample wells: 10 µL of sample + 40 µL of Sample Diluent.
- Blank well(s): Leave empty.
- Add Detection Antibody: Add 100 µL of HRP-conjugated detection antibody to all wells except blank. Cover with plate sealer and incubate at 37°C for 60 minutes.
- Wash: Aspirate and blot dry. Fill each well with 1× Wash Buffer, let stand for 1 minute, then discard. Repeat for a total of 5 washes. An automated plate washer (350 µL/well, 1-min soak) may be used.
- Substrate: Add 50 µL each of Substrate A and Substrate B. Incubate in the dark at 37°C for 15 minutes.
- Stop & Read: Add 50 µL Stop Solution. Measure absorbance at 450 nm within 15 minutes.
Total hands-on time: approximately 2 hours.
Materials Required but Not Supplied:
- Microplate reader (450 nm)
- Adjustable pipettes and tips (0.5–1000 µL)
- 37°C incubator or water bath
- Distilled/deionized water
- Absorbent paper, wash bottle or automated plate washer
- Protease inhibitor cocktail (for tissue homogenates)
- 1% H₂O₂ (optional, for high-peroxidase tissues)
Throughput
| Format | Max. Number of Samples (Single Wells) |
|---|---|
| 48-well | 39–40 samples |
| 96-well | 85–88 samples |
Includes 6 standard wells and 1–2 blank wells. For duplicate measurements, divide the sample number accordingly.
Expand Your Equine Inflammation Panel
For comprehensive profiling of equine inflammatory mediators, pair this horse IL-8 kit with:
- Horse Interleukin-6(IL-6) ELISA Kit – For quantitative measurement of interleukin-6 in equine serum, plasma, cell culture supernatant, and tissue homogenates.
- Horse Interleukin-1β (IL-1β) ELISA Kit – a master pro‑inflammatory cytokine elevated in equine septic arthritis, osteoarthritis, and acute laminitis.
- Horse PTH ELISA Kit – Equine Parathyroid Hormone Quantitative Assay for Calcium Metabolism Research
- Horse Interferon γ (IFN-γ) ELISA Kit – High-Sensitivity Sandwich Assay for Th1 Immune Response
Contact Us to explore our full range of equine ELISA kits covering TNF‑α, IL‑6, IL‑10, growth factors, and acute‑phase proteins to build a complete cytokine and chemokine profiling platform.
Frequently Asked Questions
Q: Why do I need to multiply my result by 5?
A: The assay protocol dilutes the sample 5-fold in the well (10 µL sample + 40 µL diluent). The standard curve is processed identically, so the interpolated value reflects the diluted concentration. Multiply by 5 to obtain the actual concentration in the original sample.
Q: Can this kit detect IL-8 from other species?
A: No. Extensive testing has confirmed no cross-reactivity with human, mouse, rat, bovine, or canine IL-8. The kit is validated solely for equine samples.
Q: How should I prepare tissue homogenates to ensure accuracy?
A: Always use fresh protease inhibitors in ice-cold buffer, homogenize on ice, and incubate for 30 min before centrifugation. For liver, kidney, or other high-peroxidase tissues, treat the supernatant with 1% H₂O₂ for 15 min prior to the assay to prevent false-positive signals.
Q: What is the normal IL-8 concentration in horse serum?
A: IL-8 levels in healthy horses are typically very low and may be near or below the detection limit. We strongly recommend running pilot experiments with your specific population to determine baseline ranges.
Q: My sample reads above 240 pg/mL after applying the 5× factor. What should I do?
A: Pre-dilute the sample further with the provided Sample Diluent (e.g., an additional 1:2 or 1:5), re-assay, and multiply by both the 5× in-well dilution and the extra dilution factor.
Q: Can I use hemolyzed or lipemic samples?
A: Hemolyzed samples may interfere with the assay. We recommend collecting fresh samples and avoiding hemolysis. Lipemic samples should be centrifuged thoroughly; if possible, use a lipid-clearing agent or collect non-lipemic samples.
Ordering & Contact Information
| Item | Details |
|---|---|
| Catalog Number | Yanda-HORSE-IL-8 |
| Formats | 48-well / 96-well |
| Package Dimensions | 16 cm × 12.5 cm × 9.5 cm (neutral white box) |
| Shipping | Cold-chain transport (2–8°C), protected from light and moisture |
| Storage | 2–8°C; do not freeze |
| Shelf Life | 6 months |
| Intended Use | For research use only, not for diagnostic procedures |
Contact us
- WhatsApp: +86 198 3186 0669
- Email: songhawyble@gmail.com
For pricing, bulk orders, or customization inquiries, please reach out via the above channels.
Technical Documents & Support
- Instruction Manual – Available for download [insert link] or upon request.
- MSDS – Supplied with the kit or available on request.
- Technical Support – For validation data, protocol assistance, or troubleshooting, contact us via WhatsApp or email.
Disclaimer
This product is for laboratory research use only and is not approved for clinical diagnosis, treatment, or prognosis in humans or animals. The manufacturer assumes no responsibility for consequences arising from improper use, protocol deviations, or unauthorized applications. Users are responsible for validating assay performance in their own sample matrices and experimental conditions.research with precision, convenience, and complete data transparency.
