Many dyes have varying degrees of affinity for DNA and can thus be used for chromosome staining. In routine chromosome staining, dyes such as Giemsa, orecin, Feulgen, and carbol fuchsin generally yield satisfactory results. Thionine can be used for chromosome staining in samples such as oral mucosa, urine, amniotic fluid, chorionic villus cells, and artificially cultured cells. It is particularly suitable for staining sex chromatin, which is often difficult to stain. Additionally, thionine can be used to stain substances such as Nissl bodies and mast cells.
Thionine Staining Solution (1%) is primarily composed of phosphate salts and thionine. When staining Nissl bodies, they appear deep blue, while mast cells appear purplish-red. This reagent is intended for research use only and is not suitable for clinical diagnosis or other purposes.
Reagents, consumables and Equipments not provided
- PBS or physiological saline, distilled water, 1 mol/L HCl, fixative solution (methanol:glacial acetic acid = 3:1)
- Glass slides, cover slips
- Microscope
Instructions for Use (For Reference Only)
I. Sample Processing
Oral Mucosal Cells
(1) Rinse the mouth several times with PBS or physiological saline to remove as many bacteria and debris as possible.
(2) The operator should hold the patient’s lower lip with one hand and use the blunt end of a tongue depressor or toothpick to scrape the mucosa from the inner cheek or lower lip. Discard the first scraping.
(3) Scrape the same area several times consecutively and rinse the scraped material into a centrifuge tube containing 5 mL of physiological saline.
(4) Centrifuge at 1500 × g for 10–15 minutes, discard the supernatant, and retain the cell pellet.
(5) Add 10 mL of fresh fixative solution (methanol:glacial acetic acid = 3:1), gently mix to form a suspension, and let stand at room temperature for 30 minutes.
(6) Place one drop of the suspension onto a pre-cooled, clean glass slide and air dry.
Exfoliated Cells in Urine
(1) Collect a clean midstream urine sample from the patient, mix well, and transfer 10 mL to a centrifuge tube.
(2) Centrifuge at 1500–2000 × g for 10–15 minutes, discard the supernatant, and retain the cell pellet.
(3) Add 10 mL of fresh fixative solution (methanol:glacial acetic acid = 3:1), gently mix to form a suspension, and let stand at room temperature for 30 minutes.
(4) Centrifuge at 1500 × g for 10–15 minutes, discard the supernatant, and retain the cell pellet.
(5) Depending on the number of cells, add a few drops of freshly prepared fixative solution and mix thoroughly to form a suspension.
(6) Place one drop of the suspension onto a pre-cooled, clean glass slide and air dry.
Amniotic Fluid Cells
(1) Using routine gynecological procedures, transabdominally aspirate approximately 10 mL of amniotic fluid from a pregnant woman at around 16 weeks of gestation into a centrifuge tube. Discard the first 2–3 mL of amniotic fluid to avoid contamination with maternal cells.
(2) Centrifuge at 1500 × g for 10 minutes, discard the supernatant, and retain the cell pellet.
(3) Add 10 mL of fresh fixative solution (methanol:glacial acetic acid = 3:1), gently mix to form a suspension, and let stand at room temperature for 30 minutes.
(4) Centrifuge at 1500 × g for 15 minutes, discard the supernatant, and retain the cell pellet.
(5) Depending on the number of cells, add a few drops of freshly prepared fixative solution and mix thoroughly to form a suspension.
(6) Place one drop of the suspension onto a pre-cooled, clean glass slide and air dry.
II. Chromatin Staining
Place the slide specimen in 1 mol/L HCl and incubate at 37°C for 20 minutes.
Rinse thoroughly with distilled water and air dry naturally.
Immerse the slide specimen in Thionine Staining Solution (1%) and stain for 15–20 minutes.
Rinse with distilled water and air dry naturally.
Under low magnification, locate evenly dispersed cell clusters, then switch to oil immersion for detailed observation.
Staining Results
Metachromatic substances appear purplish-red, while other structures appear blue.
Precautions
1. During centrifugation to obtain cell pellets, for certain cell types, if the pellet is insufficient, the centrifugation force or time may be appropriately increased.
2. For your safety and health, please wear a lab coat and disposable gloves during operation.
3. This product is for research use only. Any other use is strictly prohibited.
