slides to be stained for
1: appendix, 2-3:
gastrointestinal stromal tumours, 4: breast 'myeloid
sarcoma' (acute myeloid leukaemia),
Criteria for assessing a CD34 staining as
optimal were: a strong and distinct cytoplasmic reaction with
membrane accentuation in the endothelial cells of small vessels in the appendix and
the liver (portal tracts and zone 1 sinusoids),
tumour cells of the GISTs, and a high proportion of tumour
cells in the myeloid sarcoma,
without any staining of epithelial and smooth muscle cells.
64 laboratories submitted stainings. At the assessment 29 laboratories achieved optimal staining (45%), 19
good (30%), 9 borderline (14%), and 7 poor staining (11%).
57 used mAb QBend10 (DakoCytomation (25), Novocastra (14),
Immunotech (6), Neomarkers (4), Ventana (2), Monosan (2),
BioGenex (2) , Cell Marque (1) and Bio-Zac (1)); 6 used mAb
My10 (Becton Dickinson), and one used mAb BI-3C5 (Zymed).
Optimal stainings could
be achieved with both mAb QBend10 and mAb My10. The dilution appeared to be highly dependent
of origin of the Ab.
Mandatory for an optimal
result was HIER.
Most optimal protocols used Tris-EDTA/EGTA pH 9, but also
citrate pH6 was used in combination with a sensitive
visualization system. Laboratories using proteolytic
pre-treatment or no pre-treatment obtained good (but not
optimal) results in some cases. While it was possible in
most protocols to obtain a proper staining of endothelial
cells in large vessels, staining of small vessel endothelium,
one of the GISTs and the acute myeloid leukaemia provided an
The most frequent causes of insufficient stainings (often
in combination) were:
too low primary
(too low pH and/or too short heating time)
- A less sensitive visualization system