Luxol Fast Blue Staining Solution

Guidelines (The following is our recommended protocol, which serves only as a guide and should be modified according to your specific needs).
(1) Kidney Tissue Staining: Combined LFB + PAS Method
1. Applicable tissues: Rat kidneys (fixed in Regaud's solution, 5 μm paraffin sections).
2. Deparaffinize sections, then rehydrate through absolute ethanol and 95% ethanol.
3. Immerse sections in 0.1% Luxol Fast Blue solution (dilute this product to 0.1% with 95% ethanol) and incubate at 55°C for 2-4 h.
4. Wash away excess stain with 95% ethanol, then rinse with distilled water.
5. Differentiate with 0.05% lithium carbonate aqueous solution for 3-5 seconds, followed by three rinses with 70% ethanol, and then rinse with distilled water.
6. Treat with 0.5% periodic acid for 5 min, then rinse with running water.
Stain with Schiff reagent for 20 min.
7. Rinse three times with sulfite solution, 1-2 min each time, then rinse with running water.
8. Dehydrate, clear, and mount sections with resin.
9. Staining results: Brush borders appear deep red, mitochondria appear dark blue, and hyaline droplets and basement membranes appear purple.
(2) Kidney/Liver/Testis Tissue Staining: LFB + Phloxine Mitochondrial Staining Method
1. Applicable tissues: Kidneys, livers, and testes of mice and bullfrogs (fixed in formalin-Zenker or Regaud's solution, 2-5 μm paraffin sections).
2. Deparaffinize sections, then rehydrate through absolute ethanol.
3. Immerse sections in 0.1% Luxol Fast Blue solution (containing 0.05% glacial acetic acid) and incubate at 58°C for 3-6 h.
4. Wash away excess stain with 95% ethanol, then rinse with distilled water.
5. Differentiate with 0.05% lithium carbonate aqueous solution for approximately 10 seconds, followed by three rinses with 70% ethanol until partial areas become decolorized and transparent.
6. Rinse with distilled water.
7. Counterstain with 0.5% Phloxine aqueous solution for 2-3 min.
8. Differentiate with 5% phosphotungstic acid aqueous solution for 2 min, then rinse with running water for 5 min.
9. Dehydrate with ethanol, clear, and mount sections with resin.
10. Staining results: Mitochondria appear bright green, cell nuclei appear red, nucleoli appear green, and cytoplasm appears pink to pale green; the middle piece of sperm (area with dense mitochondria) appears dark green.
(3) Universal Tissue Sections: LFB + FeCl₃ Phase-Contrast Microscopy Staining Method
1. Applicable tissues: Paraffin sections (4-5 μm) fixed with various fixatives (Bouin's, Helly's, Zenker's, Heidenhain's SUSA, etc.).
2. Deparaffinize sections, then rehydrate to 70% ethanol.
3. (If mercury-containing fixative is used) Remove mercury with Lugol's solution → treat with 5% sodium thiosulfate → rinse with water → return to 70% ethanol.
4. Immerse sections in 1% Luxol Fast Blue working solution (mix this product with FeCl₃-hydrochloric acid ethanol solution at an equal ratio, prepare immediately before use) and stain at room temperature for 1.5 h.
5. Rinse with 70% ethanol.
6. Dehydrate, clear, and mount sections.
7. Staining results: Under a phase-contrast microscope (dark-phase objective), cell membranes, nuclear membranes, chromatin, and cytoplasmic structures appear black against a green background; structures such as terminal bars, epithelial tonofibrils, brush borders, striated borders, zymogen granules, and filamentous mitochondria in renal tubules can be clearly visualized.
8. Note: Sections stained by this method are only suitable for observation under a phase-contrast microscope, not for observation under a regular bright-field microscope.
Luxol Fast Blue Staining Solution selectively stains paraffin sections of mouse/bullfrog liver, kidney, and testis tissues to display blue-green mitochondria and basement membranes^[1][2].
Luxol Fast Blue Staining Solution (0.1% w/v; 56-58℃; single application; 3-16 h) stains paraffin/frozen sections of brain and spinal cord from model mice or rats with conditions such as ICH and SCI, enabling visualization of myelin loss and demyelination severity^[1][2][3].