Hematoxylin-Eosin Staining Kit (Including Differentiation and Bluing Solutions)
Based on 1 Customer Validation
MCE Hematoxylin-Eosin Staining Kit (Including Differentiation and Bluing Solutions) contains staining solutions, differentiation solution, and bluing solution. Through optimized formulations, it helps to adjust the staining effect, ensuring proper staining intensity and further improving experimental accuracy.
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Storage :
Room temperature, 1 year. Protected from light.
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Description & Advantages
Hematoxylin is a natural basic dye extracted from Haematoxylon campechianum that binds to negatively charged acidic substances, such as DNA, staining the chromatin in cell nuclei blue-purple. Eosin is a synthetic acidic dye that binds to positively charged amino cations in proteins, staining the cytoplasm and extracellular matrix red or pink. After staining with Hematoxylin and Eosin, the cell nuclei appear blue or blue-purple, while the cytoplasm and extracellular matrix are stained red or pink. This significantly enhances the contrast of tissue structures, making histological studies more intuitive.
MCE Hematoxylin-Eosin Staining Kit (Including Differentiation and Bluing Solutions) contains staining solutions, differentiation solution, and bluing solution. Through optimized formulations, it helps to adjust the staining effect, ensuring proper staining intensity and further improving experimental accuracy.
This kit is suitable for staining tissue paraffin sections, frozen sections, and cultured cells in immunohistochemistry, particularly for histological and pathological studies where precise observation of cell nuclei and cytoplasm structures is required.
Protocol
Reagent Preparation
Fixative: 4% Paraformaldehyde Fixative Solution. Other reagents required: Xylene, neutral balsam or mounting medium, absolute ethanol, 95% ethanol, 90% ethanol, 80% ethanol, 70% ethanol, distilled water, etc.
Staining of Paraffin Sections
1. Deparaffinization: Treat sections with xylene for 5-10 min, then replace with fresh xylene and treat for another 5-10 min (deparaffinization should be as complete as possible; if the temperature is low, this can be performed in an incubator at 60-70°C). Optional step: treat with absolute ethanol 2 times, 3-5 min each. Then treat sequentially with 95%, 90%, and 80% ethanol for 3-5 min each, and finally rinse with distilled water (tap water or 30-40°C warm water can also be used) for 1-3 min.
2. Staining: Stain with Hematoxylin Staining Solution for 3-8 min, rinse with distilled water for 5-10 s; treat with Ethanol Differentiation Solution for 2-5 s, rinse with distilled water or tap water for 20-30 s; treat with Bluing Solution or warm water for 20-40 s to blue the sections, rinse with distilled water or tap water for 30-60 s; stain with Eosin Staining Solution for 20-60 s, rinse with distilled water or tap water for 30-60 s.
3. Dehydration, Clearing, and Mounting: Treat sequentially with 80% ethanol for 10-20 s, 90% ethanol for 10-20 s, 95% ethanol 2 times (1-2 min each), and absolute ethanol 2 times (2-3 min each); clear with xylene 3 times, 2-3 min each; mount with neutral balsam or mounting medium and observe under a microscope.
Staining of Frozen Sections
1. Fixation: (Optional) Treat with ether-ethanol mixed fixative for 5-10 s (this fixative is prepared by mixing ether and 95% ethanol in equal volumes with the addition of an appropriate amount of acetic acid, and stored sealed); rinse with distilled water or tap water for 2-5 s.
2. Staining: Stain by dropping Hematoxylin Staining Solution onto the sections for 1-5 min (can be heated to 50°C), rinse with distilled water or tap water for 2-5 s; treat with Ethanol Differentiation Solution for 2-5 s, rinse with distilled water or tap water for 2-5 s; treat with Bluing Solution or warm water for 2-5 s to blue the sections, rinse with distilled water or tap water for 5-10 s; stain with Eosin Staining Solution for 2-20 s, rinse with distilled water or tap water for 5-10 s.
3. Dehydration, Clearing, and Mounting: Treat sequentially with 80% ethanol for 1-2 s, 95% ethanol for 1-2 s, and absolute ethanol for 2-5 s; (optional) treat with phenol-xylene (1:3) for 2-5 s; clear with xylene 3 times, 2-5 s each; mount with neutral balsam or mounting medium and observe under a microscope.
Staining of Cell Samples
1. Fix with 4% paraformaldehyde for 10-20 min.
2. Rinse with tap water 2 times, 2 min each; then rinse with distilled water 2 times, 2 min each.
3. Follow the same staining, dehydration, clearing, and mounting steps as described above for paraffin sections; treatment times can be shortened as appropriate.
Interpretation of Staining Results
Cell nuclei appear blue; cytoplasm, muscle fibers, collagen fibers, and thyroid colloid, etc., appear in varying shades of red; keratin, red blood cells, etc., appear bright orange-red.
Note: The differentiation step after staining can improve the clarity of nuclear and cytoplasmic staining, but should not be prolonged excessively; the differentiation time with ethanol can be adjusted according to section thickness, tissue type, and the freshness of the differentiation solution. Rinsing after differentiation is intended to thoroughly remove residual acid. The staining solution can be reused and should only be replaced when staining performance is noticeably reduced. Ethanol is volatile, so ethanol solutions of various concentrations should preferably be prepared fresh before use. At lower temperatures, hematoxylin may stain less readily, so staining time can be extended accordingly; at higher room temperatures, deparaffinization time can be shortened. The Bluing Solution can also be substituted with 0.2-1% ammonia water, Scott's tap water substitute, or 0.1-1% lithium carbonate solution. Fluorescent staining, such as immunofluorescence, is not recommended following staining with this kit.
Storage
Room temperature, 1 year.
Protected from light.
Attention
1. Differentiation after staining helps to clarify the nuclear-cytoplasmic staining, but the time should not be too long. The differentiation time with ethanol can be adjusted based on the section thickness, tissue type, and the freshness of the differentiation solution. The rinse after differentiation is to thoroughly remove the acid.
2. Staining solution can be reused; replace with fresh staining solution when staining performance significantly decreases.
3. Since ethanol is highly volatile, graded ethanol solutions should be freshly prepared before use to ensure consistent dehydration performance.
4. Hematoxylin staining may not perform well at lower temperatures; it is advisable to extend the staining time appropriately.
5. At higher room temperatures, the deparaffinization time can be reduced slightly.
6. Bluing solution can also be replaced with 0.2-1% ammonia solution, Scott’s bluing solution, or 0.1-1% lithium carbonate solution.
7. After staining with this kit, subsequent immunofluorescence or other fluorescence-based staining is not recommended, as it may interfere with fluorescence signals or the interpretation of staining results.
8. This product is for R&D use only, not for drug, household, or other uses.
9. For your safety and health, please wear a lab coat and disposable gloves to operate.
Components
| Components | HY-K0315A-400 mL | HY-K0315A-2000 mL |
|---|---|---|
| Hematoxylin Staining Solution | 100 mL | 500 mL |
| Ethanol Differentiation Solution (Acidic) | 100 mL | 500 mL |
| Bluing Solution | 100 mL | 500 mL |
| Eosin Staining Solution | 100 mL | 500 mL |
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