Concerning staining of Entamoeba histolytica:

A PAS with any counterstain (light) would work. We've done
PAS-Hematoxylin,
doing about a 30 second hematoxylin. Works great.

Metanil yellow, commonly used as a counterstain for mucicarmine, would
work also. Stain for about 30 seconds.

Metanil yellow - 0.25 g
Distilled water - 100 mL
Acetic acid - 0.25 mL

Dissolve together. Good for about 2 weeks. May be reused.

The only Martius yellow counterstain I've ever used was with the
MSB stain for fibrin. It stains everything yellow, especially
RBC's. Entamoeba histolytica can ingest RBC's, so this
counterstain would help demonstrate the RBC's. 

(In the MSB stain, eventually, only the RBC's are yellow. MSB 
stain is similar to Masson Trichrome with the addition of 
Martius yellow for the RBC's. It's a quatrachrome stain, I guess.)

The following is the procedure used in the MSB stain.
If I were to guess how to use it for the Amoeba stain, after 
staining in PAS with water washing, rinse in 95% alcohol 
for about 30 seconds. Stain in martius yellow for 30 seconds.
Rinse 5-10 seconds in 95% alcohol, then in abs. alcohol. 
Clear. Coverslip.

Martius yellow (acid yellow 44) - 0.5 g
95% alcohol - 100 mL
Phosphotungstic acid - 2.0 g

Dissolve martius yellow in alcohol. Then dissolve in phosphotungstic
acid. Good for several months. May be reused.

As for a control: This is not a stain for all amoebas, but usually
Entamoeba histolytica, which causes amoebic dysentery and amoebic
colon ulcers, but can burrow through the intestine wall, into a 
blood vessel, and end up in other parts of the body, commonly 
as liver abscesses. E. histolytica has a glycogen store in it. 
So if you can't get a control with E. histolytica in it, a 
glycogen control would be OK, but obviously not the best.

Hope this helps.

**********************
Peggy A. Wenk, HTL(ASCP)
William Beaumont Hospital
Royal Oak, MI 48073
SEENRD@aol.com wrote:
> 
> has anyone out there ever done a PAS with a martius yellow counterstain for
> amoeba? if so, what percentage of martius yellow did you use and how long did
> you counterstain? one more question.....are there any good sources for amoeba
> controls? thanks in advance for the info!!
> 
>    donna