Giemsa Staining Solution (Chromosome G-Banding)
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MCE Giemsa Stain Solution for Chromosome G-Banding is a concentrated staining solution specifically formulated for chromosome G-banding and cytogenetic karyotype analysis. It is composed primarily of Giemsa dye and methanol and contains specialized counterstaining components. Before use, dilute the stain with buffer at a stain-to-buffer ratio of 1:10. The resulting working solution is designed to provide clear, uniform staining for standardized cytogenetic workflows.
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Storage :RT, 2 years. Protect from light.
- Manual (3239KB)
- COA
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Description & Advantages
Giemsa dye is composed of Azure II and eosin. Its staining principle is based on differences in the affinity of cellular components for acidic and basic dyes. After controlled trypsin digestion, staining chromosome preparations with this product produces transverse bands of varying intensity and width along the length of each chromosome. The 22 pairs of human autosomes and the X and Y chromosomes have relatively characteristic banding patterns, enabling chromosome identification, pairing, and karyotype analysis. In addition to chromosome G-banding, this reagent can be used to stain blood and bone marrow smears, tissue sections, bacteria, and protozoan parasites (such as Plasmodium, Trypanosoma, and Leishmania). It can also be used in combination with Wright Stain or May-Grünwald Stain to improve visualization of nuclei, cytoplasm, and related structures.
MCE Giemsa Staining Solution (for Chromosome G-Banding) is a concentrated compound stain designed for chromosome G-banding and cytogenetic karyotype analysis. The reagent is formulated primarily with Giemsa dye and methanol and contains a specialized counterstaining component. For use, the staining solution can be diluted 1:10 with buffer. The resulting working solution provides clear, uniform staining and is suitable for standardized procedures in cytogenetics laboratories.
Eosin binds to basic proteins in the cytoplasm and eosinophilic granules, producing a pink color. Azure dyes, including Azure II, bind to nuclei and other basophilic structures, producing a blue-purple color. This enhances the color contrast among nuclei, cytoplasm, granules, and other structures, facilitating morphological examination under a microscope.
Protocols
Materials Required but Not Provided
Reagents: phytohemagglutinin (PHA), colchicine, 0.075 M KCl, ice-cold fixative (glacial acetic acid:methanol = 1:3), and 0.025%–0.05% trypsin solution (Cat. No. HY-K3009, dilute with 0.9% NaCl).
Equipment and supplies: incubator, microscope slides, and microscope.
Sample Preparation
The following procedure uses peripheral blood lymphocytes as an example.
1. Inoculation and Culture
Add approximately 0.5 mL PHA (working concentration: 60 μg/mL) to a culture tube containing 3–5 mL human peripheral blood. Incubate at 37 °C in 5% CO2 for 72 h.
2. Metaphase Arrest
Add colchicine to a final concentration of 0.5 μg/mL. Continue incubation for 15–30 min.
3. Cell Collection and Hypotonic Treatment
Transfer the culture to a centrifuge tube and centrifuge at 2,000 × g for 5 min. Discard the supernatant. Add 5–10 mL of 0.075 M KCl and gently resuspend the cells. Incubate at 37 °C for 10–20 min, then centrifuge at 2,000 × g for 5 min and discard the supernatant.
4. Cell Fixation
Gently disperse the cell pellet. Add 5–10 mL of freshly prepared, ice-cold fixative (glacial acetic acid:methanol = 1:3) dropwise, mix, and incubate at 4 °C for 10 min. Centrifuge at 2,000 × g for 5 min and discard the supernatant. Add another 5–10 mL of ice-cold fixative, repeat the fixation and centrifugation steps, and discard the supernatant.
5. Slide Preparation and Drying
Gently resuspend the cell pellet in 0.5–1 mL of fresh fixative. Drop the cell suspension onto clean microscope slides and allow the slides to air-dry before staining.
Giemsa Staining
1. Prepare the Working Solution
Mix Giemsa Stain for Chromosome Analysis and Giemsa Staining Buffer at a 1:9 ratio. Mix thoroughly before use. Prepare the working solution fresh. If the staining result is unsatisfactory, optimize the dilution ratio in a preliminary experiment.
2. Trypsin Treatment
Immerse the slides in 0.025%–0.05% trypsin solution for 5–30 min. Optimize the treatment time according to the enzyme concentration and lot, temperature, and slide preparation. Immediately after treatment, rinse gently with 0.9% NaCl (w/v) to stop digestion.
3. Giemsa Staining
Apply enough Giemsa working solution to completely cover the specimen area. Stain at room temperature for 15–30 min.
4. Rinsing and Drying
Slowly add distilled water at one edge of the slide to gently rinse off excess stain without directing the stream onto the specimen area. Allow the slides to air-dry.
Storage
RT, 2 years.
Protect from light.
Attention
1. After staining, do not pour the staining solution directly off the slide or rinse the stained area with a strong stream of water. Slowly apply the rinsing solution from one end of the slide and allow the stain to flow off naturally to prevent sample detachment or uneven staining.
2. If staining is too dark, shorten the staining time or reduce the working solution concentration. If staining is too light, extend the staining time or increase the working solution concentration.
3. Buffer pH affects the staining hue and band contrast. Prepare the buffer according to the specified conditions.
4. A slight metallic sheen on the surface of the diluted Giemsa working solution is normal, but it should not be used as the sole indicator of stain performance. If a distinct surface film or precipitate forms, filter or remove it before use.
5. Prepare the staining working solution fresh before use. It may be reused briefly for samples within the same batch, but repeated reuse is not recommended because it may affect staining quality.
6. This product is intended for scientific research by trained personnel only. It must not be used for clinical diagnosis or treatment, or in food or pharmaceutical products.
7. For your safety and health, wear a lab coat and disposable gloves when handling this product.
Components
| Components | HY-K0616 -100 mL |
|---|---|
| Giemsa Staining Solution (Chromosome G-Banding) | 10 mL |
| Giemsa Staining Buffer | 100 mL |
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