Simultaneous Detection of Leukemia Blast Cells on PB/BM — Smears Combining May-Gruenwald Giemsa- and APAAP Staining
For subtyping acute leukemias the French-American-British (FAB) classification system has been the most widely used system. The system uses morphological and cytochemical criteria to initially define myeloid and lymphoid groups. Since the recent availability of monoclonal antibodies (MoAbs) recognizing various differentiation antigens the diagnostic precision has increased. Immunologic assessments of cell-surface antigen expression have now been recognized as providing an additional set of criteria upon which to base a more comprehensive description of leukemic cells.
In the present study we describe the combination of Pappenheim staining with an immunocytochemical method (APAAP-complex-technique), providing the visualization of morphological features as well as cell surface antigens, e.g., CD2, CD3, CD7, CD10, CD 13, CD 14, CD 19, CD20, CD24, CD34, HLADR and TdT in air-dried cell smears from different subtyps of leukemic blasts (ALL/AML) form children and from leukemia-derived cell lines (Molt-4, REH, U937).
Following May-Gruenwald Giemsa staining, the morphological structure of cells was examined under an optical microscope, photographed, destained, fixed and immunolabeled by a modified alkaline phosphatase anti-alkaline phosphatase (APAAP) complex technique with repeated incubation steps using Silicone-Chamber-System (SCS).
After finishing the immunoenzymatic colour reaction by the application of naphthol AS-MX-phosphate and Fast Red TR Salt as chromogenic substrate the marked cell areas were visualized and photographed again.
Performing the proposed schedule, it was still possible to classify acute leukemias according to their surface marker expression and to distinguish simultaneously between myeloid and lymphoid cells by morphological criteria.
In conclusion, the application of simultaneous staining procedure allows the direct comparison of morphology by light microscopy and the marker expression at single leukemic blast cell level.
KeywordsAcute Myeloid Leukemia Acute Leukemia Surface Marker Expression Form Child Cytospin Preparation
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