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FIXATION:        10% buffered neutral formalin.

TECHNIQUE:        Paraffin sections at 5 μm

Modified Weigerts Iron Hematoxylin

(See Gomoris One Step Trichrome Method)

Aniline-Eosin Solution

Eosin Y C.I. 45380 --------------------------------------------------------------        1.5 gm

Alcohol, 80% ---------------------------------------------------------------------        100.0 ml

Aniline -----------------------------------------------------------------------------        3.0 ml

Acetic Acid -----------------------------------------------------------------------        1.0 ml

       Dissolve the eosin Y in the distilled water and then add the alcohol,

       aniline and acetic acid.

Naphthol Green B Solution

Naphthol green B C.I. 10020 ---------------------------------------------------        -        1.0 gm

Distilled water --------------------------------------------------------------------        100.0 ml

Acetic Acid -----------------------------------------------------------------------        1.0 ml

Use control slide.

1.        Deparaffinize and hydrate to distilled water.
2.        Modified Weigerts iron hematoxylin for one minute.
3.        Wash well in running tap water.
4.        Differentiate in 0.5% acid alcohol for 10 seconds.
5.        Wash in running tap water for 5 minutes and rinse with distilled water.
6.        Aniline-eosin solution for 5 minutes.
7.        Rinse well in distilled water.
8.        Naphthol green B solution for 5 minutes.

9.        Differentiate in 95% alcohol, two changes, until erythrocytes in the section are bright rose.  Check under microscope.

10.        Dehydrate in absolute alcohols and clear in xylene, three or four changes.

11.        Mount with synthetic resin.

Amoebae ------------------------------------------------------------                blue-green

Nuclei of amoebae -------------------------------------------------                deeper blue-green

Ingested erythrocytes ----------------------------------------------        deep rose

Connective tissue ---------------------------------------------------        green

This procedure does not stain amoebae differentially, but is useful in that it demonstrates the ingested erythrocytes well.

Another stain which is useful for demonstrating amoebae is the periodic acid-Schiff with an iron hematoxylin counterstain.

Gridley, M.F.:  A stain for endamoeba histolytica in tissue sections.  Amer. J. Clin. Path. 24:243-244, 1954.