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Gladstone Institutes Histology and Light Microscopy Core
Melanin bleach Protocol
This bleach is used in conjunction with Masson’s Fontana for melanin granules. If the positive
MF slide is ‘positive’ and the bleached one is negative then this shows the pigment is melanin
Tissue:
Formalin fixed paraffin embedded tissue sectioned at 2-10m. Can also be done
on cryosections, but quality and resolution is much better with paraffin tissues. Tissue should
be baked on prior to staining as it tends to make the sample lift.
Controls:
Skin containing melanin
Solutions:
0.25% Potassium permanganate
Potassium permanganate
diH2O
1.25g
500ml
1% Oxalic acid
Oxalic acid
diH2O
5g
500ml
Procedure:
1. Dewax and rehydrate to running tap water
2. Bleach with pot permanganate
15 min
3. Wash in running tap water
10 min
4. Decolourize section with 1% oxalic acid
20sec – 1 min
5. Perform Massons’ Fontana on the bleached and unbleached sections
OR
6. 95% alchol
1 x 1 min
7. 100% ethanol
2 x 1 min
8. Xylene
3 x 1min
9. Mount in DPX (Gurr)
Results:
Nuclei
- Black
Notes: Melanins from different tissue types require different times in pot permanganate
e.g. skin melanin
10-20min
ocular melanin
2-4hrs
26 Dec 2012
Weigert’sHx
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