STAINING FOR MAST CELLS

Mast cells are proinflammatory leukocytes that reside in the connective tissues and epithelia of all organs.  Under  allergic stimuli, their intraepithelial mucosal cell population expands dramatically (1). However, mast cell populations usually go unnoticed because they do not stain with hematoxylin and eosin and they are not autofluorescent, thus evading detection in nearly all routine preparations of paraffin or plastic embedded tissues and cell cultures. Information about mast cells is thus lost to the scientific community interested in their distribution, deployment and function. At least of equal importance is the continued ignorance of the role of the mast cell in diverse biological and pathological processes because the cells are simply not made visible to the investigator. Staining for mast cells in formalin or paraformaldehyde fixed  preparations of cells and tissues is easy.

Toluidine blue staining at low pH, a procedure that just takes a few seconds, stains the granules of mast cells in connective tissues purple( 2). Wright Giemsa and Diffquik (Dade-Behring, Newark, DE)(3) stain mast cells in connective tissue blue to purple and in some locations such as the gastric mucosa and broncho- tracheal epithelium stains the intraepithelial mucosal mast cells crimson as well .The likely basis for the differential staining of mast cells residing in intraepithelial versus  connective tissue locations is attributable to the differences in the proteoglycans to which the zymogens are bound in their secretory granules– heparin in the granules of mast cells that reside in the connective tissues and chondroitins in the cells in intraepithelial locations(4). Congo red, also a metachromatic  stain like toluidine blue, stains intraepithelial mast cells orange (5). The chloroacetate esterase procedure (6) is a universal stain for mast cells, rendering them red in color regardless of the cells’ location. Chloroacetate esterase is my procedure of choice (7).

Less commonly used stains also reveal mast cells clearly. Ruthenium red and Texas red stain mast cells in connective tissues red. Alcian blue stains mast cells in connective tissues azure. Methylene blue stains those cells dark blue. Bismark brown stains them brown. Preparations using methenamine silver, such as in the Jones stain, renders all mast cells black.

Immunocytochemistry provides another approach to mast cell identification commonly used by experts in the field but not as easy for the general investigator as the staining methods referenced above.  For human tissues anti-tryptase antibodies mark intraepithelial mucosal mast cells and anti-chymase antibodies mark mast cells in connective tissues . In mice, mMCP-5, 6, and 7 are useful for identifying mast cells in connective tissues and mMCP-1 is selective for intraepithelial mucosal mast cells (8).

When one stains for mast cells while exploring other phenomena, beneficial surprises can occur (9).

(1)    D S Friend et al., J. Cell Biology 135, 279 (1996)

(2)    L Enerback, Acta. Pathol. Microbiol. Scand. 66, 303 (1966)

(3)    M Gurish et al., J. Immunol. 168, 5730 (2002)

(4)    GW Wong et al., Signal Transduction in Mast Cells & Basophils, E Razin & J Rivera eds.,Springer-Verlag, NY (1999)

(5)    DS Friend et al., J. Immunol. 160, 5537 (1998)

(6)    LD Leder, Am. J. Dermatopath. 1, 39 (1979)

(7)    DS Friend et al., J. Immunol. 165, 344 (2000)

(8)    HRP Miller et al., Immunology 65, 559 (1988)

(9)    E Gounaris et al., PNAS 104, 104,19977 (2007)                                                    

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