Perl's Prussian Blue

Classification: pigment stain

Mechanism of staining: histochemical

Purpose: stain hemosiderin


Control tissue: spleen, bone marrow

Bone Marrow



REAGENT

REAGENT

PURPOSE

MECHANISM OF STAINING

SOURCE OF ERROR

Perl’s solution

HCl: liberates the ferric ions from the protein fraction of the hemosiderin molecule 

Potassium ferrocyanide: combines with ferric ions to produce ferri-ferrocyanide

Protein with ferric iron + HCl→  Ferric iron + potassium ferrocyanide → Potassium          ferriferrocyanide

Omitted: Hemosiderin not demonstrated.

Too short: Hemosiderin will be pale.

Too long: Background staining will occur.

Neutral red

Counterstain: Demonstrates other tissue components

Nuclei & cytoplasm

 

Ionic bonding

Omitted: Collagen, RBC, muscle, cytoplasm not demonstrated.

Too short: Collagen, RBC, muscle, cytoplasm too pale.

Too long: Hemosiderin is obscured.

 

 

Special Considerations

Hemosiderin is soluble in acid. Avoid acid fixatives. Also ensure the formalin is buffered and not acidic.
Hemosiderin is an endogenous pigment. When unstained, it will be visible as a yellow or brown pigment.

A positive control is suggested.

Bone Marrow

Red: red blood cells, collagen 

Dark blue: hemosiderin


Spleen 



References

Officer B. HIML251 Lecture notes: Histochemical Stains: Perls Prussian Blue & Periodic Acid Schiff, January 14, 2009

Bone Marrow Biopsy- Methodologies. 2008. [cited 2009 April 6].

Available from
http://bonemarrowbiopsy.files.wordpress.com/2008/12/perls-prussian-blue.png? w=408&h=306