LUXOL FAST BLUE - MODIFIED KLUVER'S - MYELIN SHEATH
PURPOSE: Myelin sheath is the white fatty non living material forming an
insulating and protective sheath around nerve fibers.
PRINCIPLE: Luxol Fast Blue is the alcohol soluble counterpart of the
soluble Alcian Blue. Staining
is due to lipoproteins, and the
mechanism is one of an acid-base reaction with salt formation; the base
of the lipoprotein replaces the base of the dye
CONTROL: Normal cerebral cortex
: 10% formalin
: Cut paraffin sections
10μ. Allow to dry in a 37°C oven
EQUIPMENT: Rinse all glassware
in DI water
, coplin jars, 60°C oven.
Luxol Fast Blue Solution:
Luxol fast blue 1.0 gm
95% alcohol 100.0 ml
10% acetic acid 5.0 ml
. Solution is stable for 1
CAUTION: Flammable liquid, avoid
contact and inhalation.
0.05% Lithium Carbonate:
Lithium carbonate 0.5 gm
CAUTION: Avoid contact and inhalation.
Eosin Y Solution
0.25% Cresyl Echt Violet:
Cresyl Echt violet 0.25 gm
10% Acetic acid 1.0 ml
. Stable for 2 years
CAUTION: Avoid contact and
SAFETY: Wear gloves
, goggles and lab coat. Work in a well ventilated
area, avoid contact and inhalation.
Acetic acid; moderately toxic on ingestion. Target organ effects on
respiratory system on inhalation. Irritant to skin
, eyes and respiratory
Lithium carbonate; irritant to eyes and skin
and eye irritant. Ingestion can cause gastrointestinal
irritation, nausea, vomiting diarrhea, blindness, death. Excessive
inhalation can cause nasal and respiratory irritation.
1. Deparaffinize and hydrate to 95% alcohol.
2. Luxol fast blue solution, overnight, 60°C .
3. Rinse in 95% alcohol.
4. Rinse in distilled water
5. Lithium carbonate solution, 5 seconds.
6. 70% alcohol, two changes, 10 seconds each.
7. Wash in distilled water
8. Repeat steps 5 - 7 until there is a sharp contrast between the blue of
the white-matter and the colorless gray-matter.
9. Rinse slides
in 70% alcohol.
10. Eosin, 1 minute.
11. Rinse in distilled water
12. Cresyl violet, 1 minute.
13. Rinse in distilled water
14. Dehydrate through 95% and 100% alcohol, clear in xylene, and
Myelinated fibers blue
Sheehan D, Hrapchak B, Theory and practice of Histotechnology, 2nd
ed,1980, pp262-264, Battelle Press, Ohio
Crookham,J, Dapson,R, Hazardous Chemicals
in the Histopathology
Laboratory, 2nd ED, 1991, Anatech