Classical morphology, esterase cytochemistry, and interphase cytogenetics of peripheral blood and bone marrow smears

We used peripheral blood (PB) and bone marrow (BM) smears in the development of two methods based on cytomorphology and esterase cytochemistry in combination with fluorescence in situ hybridization (FISH). The first method involves photodocumentation of May-Grünewald-Giemsa (MGG)-stained cells, followed by destaining in methanol-acetic acid, fixation in paraformaldehyde, and digestion with protease and RNAse before FISH using alpha-satellite probes that specify chromosomes X, 7, 8, and 17. On average, two hybridization signals were seen in 94.5% of disomic BM cells. The hybridization sensitivity was found to vary, however, both among morphologically defined hematopoietic cell lineages and among differentation levels within a lineage. In the second method, an esterase staining technique was followed by the same treatment as for MGG-stained cells. The esterases and FISH signals could be simultaneously visualized and the method was found suitable for rapid screening of in situ signals in cytochemically defined granulocytes and lymphocytes but not in monocytes. The combined methods proved very useful in elucidating the clinical significance of chromosomal abnormalities seen in two cases of leukemia.