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Monkey hypersensitive C-reactive protein (hs-CRP) elisa kit instruction manual
**Monkey Hypersensitive C-Reactive Protein (hs-CRP) ELISA Kit – Instructions for Use**
**Kit Specifications:**
This ELISA kit is available in 48-well or 96-well configurations. It includes a standard dilution of 1.5 mL × 1 vial, enzyme standard reagent of 3 mL × 1 vial (for 48-well) or 6 mL × 1 vial (for 96-well). The kit is designed for research use only.
**Calculation Method:**
To determine the concentration of hs-CRP in the sample, plot the standard curve with the standard concentrations on the x-axis and the OD values on the y-axis. Alternatively, use linear regression to calculate the equation of the standard curve and input the sample OD value to determine the concentration. Multiply the result by the dilution factor to obtain the actual sample concentration.
**Kit Composition:**
- Sealing film: 2 pieces (48) / 2 pieces (96)
- Standard: 0.5 mL × 1 vial (2700 ng/L)
- Enzyme standard package: 1×48 / 1×96
- Sample diluent: 3 mL × 1 vial (48) / 6 mL × 1 vial (96)
- Developer A: 3 mL × 1 vial (48) / 6 mL × 1 vial (96)
- Chromogen B: 3 mL × 1 vial (48) / 6 mL × 1 vial (96)
- Wash solution: 3 mL × 1 vial (48) / 6 mL × 1 vial (96)
- Concentrated wash solution: 20 mL × 20 times × 1 bottle (48) / 20 mL × 30 times × 1 bottle (96)
**Principle of Operation:**
The kit uses a double-antibody sandwich method to detect monkey hypersensitive C-reactive protein (hs-CRP). The microplate is pre-coated with a specific antibody against hs-CRP. After incubation with the sample, the HRP-labeled antibody binds to the antigen, forming an immune complex. TMB substrate is added, and the color change is measured at 450 nm. The intensity of the color is directly proportional to the concentration of hs-CRP in the sample.
**Purpose:**
This ELISA kit is intended for the quantitative determination of hypersensitive C-reactive protein (hs-CRP) in monkey serum, plasma, urine, cell culture supernatants, and other biological fluids.
**Storage Conditions & Expiration Date:**
- Storage: 2–8°C
- Shelf Life: 6 months from the date of manufacture
**Sample Preparation & Requirements:**
1. **Serum:** Allow blood to clot at room temperature, centrifuge at 2000–3000 rpm for 10–20 minutes, and collect the supernatant.
2. **Plasma:** Use EDTA or sodium citrate as anticoagulant, mix well, centrifuge, and collect supernatant.
3. **Urine:** Centrifuge at 2000–3000 rpm for 20 minutes and collect the supernatant.
4. **Cell Culture Supernatant:** Centrifuge and collect the supernatant; for intracellular components, lyse cells via freeze-thaw cycles before centrifuging.
5. **Tissue Samples:** Homogenize tissue in PBS, centrifuge, and collect the supernatant.
6. **General Note:** Process samples immediately after collection. If not tested right away, store at -20°C. Avoid repeated freezing and thawing. Do not use samples containing NaN3, as it inhibits HRP activity.
**Operation Steps:**
1. **Standard Dilution & Loading:** Prepare a serial dilution of the standard and load into designated wells.
2. **Sample Loading:** Add 40 μL of sample diluent and 10 μL of the sample to each test well.
3. **Incubation:** Seal the plate and incubate at 37°C for 30 minutes.
4. **Washing:** Wash the plate 5 times with diluted washing solution.
5. **Enzyme Reagent Addition:** Add 50 μL of enzyme-labeled reagent to each well except blank wells.
6. **Second Incubation:** Repeat incubation 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 terminate the reaction.
9. **Measurement:** Read OD values at 450 nm within 15 minutes.
**Notes:**
- Allow the kit to reach room temperature before use.
- The concentrated wash solution may crystallize; heat gently if needed.
- Use separate pipettes for each step to avoid cross-contamination.
- Always prepare a standard curve and perform duplicate measurements.
- Keep all reagents away from light.
- Follow instructions carefully and rely on microplate reader results.
- Treat all waste materials as biohazardous.
- Do not mix reagents from different batches.
- In case of conflict, the English manual takes precedence.
**Kit Performance:**
- Correlation coefficient (R) ≥ 0.95
- Intra-assay CV < 9%, Inter-assay CV < 11%
- Detection range: 0.2 IU/L – 6 IU/L
This kit provides a reliable and accurate method for measuring hs-CRP levels in monkey samples, ensuring high-quality data for research purposes.