**Human CXCL16 ELISA Kit – For the Quantitative In Vitro Determination of Human CXC-chemokine Ligand 16 Concentrations in Serum, Plasma, Cerebrospinal Fluid, Tissue Homogenate, and Body Fluids**
*For Laboratory Research Use Only. Not for Diagnostic or Therapeutic Procedures.*
This ELISA kit is designed for the quantitative measurement of Human CXCL16 in various biological samples. The method is based on a sandwich immunoassay principle, where the target protein (CXCL16) is captured by specific antibodies immobilized on a microplate. A horseradish peroxidase (HRP)-conjugated secondary antibody then binds to the complex, and a chromogenic substrate is used to develop a colorimetric signal. The intensity of the color is directly proportional to the concentration of CXCL16 in the sample.
The kit includes a set of six standard concentrations (20, 10, 5, 2.5, 1.25, and 0.625 ng/mL) that allow for the creation of a standard curve. This curve is essential for accurately determining the concentration of CXCL16 in unknown samples. The optical density (OD) values are measured at 450 nm using a microplate reader, and results are calculated by comparing the OD of the samples to the standard curve.
**Sample Collection and Storage**
- **Serum**: Use serum separator tubes. Allow samples to clot for 2 hours at room temperature or overnight at 4°C before centrifugation at 2000×g for 20 minutes. Store at -20°C. Avoid repeated freeze-thaw cycles.
- **Plasma**: Collect in EDTA or citrate tubes. Centrifuge immediately or store at -20°C.
- **Cerebrospinal Fluid (CSF)**: Centrifuge to remove particulates and store at -20°C.
- **Tissue Homogenate**: Prepare fresh or store at -20°C. Ensure no hemolysis or contamination.
- **Cell Culture Supernatants**: Centrifuge to remove debris and use immediately or store at -20°C.
**Materials Required (Not Supplied)**
- Incubator (37°C)
- Microplate reader (450 nm)
- Pipettes, tips, and absorbent paper
- Distilled or deionized water
**Reagents Provided (Store at 2–8°C)**
- 96-well microplate strips (12×8 or 12×4)
- Standard (6 vials, 0.5 mL/vial)
- Sample Diluent (6.0 mL or 3.0 mL)
- HRP-Conjugate Reagent (10.0 mL or 5.0 mL)
- 20X Wash Solution (25 mL or 15 mL)
- Chromogen A (6.0 mL or 3.0 mL)
- Chromogen B (6.0 mL or 3.0 mL)
- Stop Solution (6.0 mL or 3.0 mL)
- Closure Membrane (2 pieces)
- User Manual (1 copy)
- Sealed Bags (1 each)
**Important Notes**
- All reagents must be brought to room temperature (20–25°C) before use.
- Do not thaw samples or reagents using water baths.
- Do not use reagents beyond their expiration date.
- Use only deionized or distilled water for dilutions.
- Keep microplates sealed until ready for use. Unused strips should be stored with desiccant.
- Always use fresh pipette tips to prevent cross-contamination.
- Handle plasma as potentially infectious. Wear gloves during the procedure.
- Waste should be inactivated for at least 30 minutes before disposal.
- Substrate solutions must be used within 30 minutes of preparation.
- Chromogen B contains 20% acetone—keep away from heat and flame.
**Preparation of 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 and standards. Add 50 μL of standard or sample in duplicate to the microplate.
2. Add 100 μL of HRP-conjugate reagent to each well except the blank. Cover and incubate for 60 minutes at 37°C.
3. Wash the plate four times manually or using an automated washer.
4. Add 50 μL of Chromogen A and 50 μL of Chromogen B to each well. Incubate for 15 minutes at 37°C, protected from light.
5. Add 50 μL of Stop Solution to each well. The color will change from blue to yellow.
6. Measure OD at 450 nm. Generate a standard curve using the average OD values of the standards.
**Interpretation of Results**
- Calculate the mean OD for each standard and sample. Subtract the blank OD value from all readings.
- Plot the standard curve (OD vs. concentration). Determine the CXCL16 concentration in each sample by interpolating its OD value on the curve.
- Intra-assay and inter-assay CVs are less than 15%.
- Assay range: 0.625–20 ng/mL.
- Sensitivity: <0.1 ng/mL.
- Cross-reactivity: No significant interference observed.
**Storage**
- Store unused kit components at 2–8°C for short-term use.
- For long-term storage, keep at -20°C for up to six months.
**Safety and Disposal**
- Follow all biohazard guidelines when handling biological samples.
- Dispose of waste according to local regulations after inactivation.
This kit is ideal for researchers studying inflammatory processes, immune responses, and related pathologies. Always read the full user manual before starting the assay.
Ningbo Honor Imp & Exp Co.,Ltd , https://www.honormaket.com