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Bowl cutter series:
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High speed bowl cutter 125L
High speed bowl cutter 200L
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Mixer series
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The application of Chopper And Mixer Series
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Smoked sausage
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The vacuum mixer adopts double shaft and helix paddle structure, which will mix raw material in circle movement, to reach the perfect mixing effect.
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**Human Thrombomodulin (TM) ELISA Kit – Instructions for Use**
This reagent is intended solely for research purposes.
**Purpose:**
The Human Thrombomodulin (TM) ELISA Kit is designed to quantitatively determine the concentration of thrombomodulin in human serum, plasma, and other biological fluids.
**Principle of Operation:**
This assay employs a sandwich ELISA method. A microplate pre-coated with anti-TM antibodies is used as the solid phase. The sample is added, followed by a biotinylated detection antibody. Horseradish peroxidase (HRP)-labeled secondary antibody then binds to the complex. After washing, TMB substrate is added, which turns blue in the presence of HRP and changes to yellow upon addition of an acidic stop solution. The intensity of the color is directly proportional to the TM concentration in the sample. Absorbance is measured at 450 nm using a microplate reader, and concentrations are calculated from a standard curve.
**Kit Components:**
- 48-well or 96-well configuration
- Coated microplate (1 × 48 or 1 × 96)
- Standard (36 μg/L, 0.5 mL × 1 bottle)
- Standard diluent (1.5 mL × 1 bottle)
- Enzyme-labeled reagent (3 mL × 1 bottle)
- Sample diluent (3 mL × 1 bottle)
- TMB substrate A and B (3 mL × 1 bottle each)
- Wash buffer (20× concentrate, 20 mL × 20 times or 30 times)
- Sealing film (2 pieces)
- Storage: 2–8°C
**Sample Preparation:**
- **Serum:** Allow blood to clot at room temperature for 10–20 minutes, centrifuge at 2000–3000 rpm for 20 minutes. Collect supernatant; if precipitated, re-centrifuge.
- **Plasma:** Use EDTA or sodium citrate as anticoagulant. Mix and centrifuge as above.
- **Urine:** Centrifuge at 2000–3000 rpm for 20 minutes. Collect supernatant.
- **Cell culture supernatant:** Centrifuge after collection. For intracellular components, lyse cells via freeze-thaw cycles, centrifuge, and collect supernatant.
- **Tissue:** Homogenize in PBS, centrifuge, and collect supernatant. Store samples at 2–8°C after thawing.
**Storage & Handling:**
- Keep all components refrigerated at 2–8°C.
- Avoid repeated freezing and thawing.
- Do not use samples containing NaN₃, as it inhibits HRP activity.
**Procedure Summary:**
1. Prepare standard dilutions in a serial manner.
2. Add 40 µL sample diluent and 10 µL sample to each well.
3. Seal and incubate at 37°C for 30 minutes.
4. Wash 5 times with diluted wash buffer.
5. Add 50 µL enzyme-labeled reagent to each well.
6. Incubate again at 37°C for 30 minutes.
7. Add TMB substrate A and B, incubate for 15 minutes.
8. Stop reaction with 50 µL stop solution.
9. Measure OD at 450 nm within 15 minutes.
**Notes:**
- Equilibrate kit at room temperature before use.
- Check pipette accuracy; use a multichannel pipette for high-throughput.
- Always include blank wells and prepare a standard curve.
- Avoid cross-contamination by using a new sealing film for each test.
- Store substrates away from light.
- Follow instructions precisely.
**Data Analysis:**
Plot standard curve with TM concentration vs. OD values. Use linear regression to calculate sample concentration. Multiply by dilution factor for accurate results.
**Performance:**
- Intra- and inter-assay variability <9% and <15%, respectively.
- Linear range: 1–30 µg/L.
- Correlation coefficient R ≥ 0.92.
**Storage & Shelf Life:**
- Store at 2–8°C.
- Valid for 6 months from date of manufacture.
This kit provides a reliable and efficient method for detecting thrombomodulin levels in various biological samples. Proper handling and adherence to protocol ensure accurate and reproducible results.