Cytogenetic analysis of cells from bone marrow aspirates has become a valuable tool in the subclassification of acute non-lymphocytic leukemias. Yet the standard techniques utilized for the visualization of leukemic chromosomes are often not adequate for detailed analysis and chromosomal abnormalities can be overlooked with the relatively few and contracted metaphases obtained (250-300 bands per haploid set). With the amethopterin cell synchronization technique developed in this laboratory, a relatively large number of cells in prometaphase (850 band stage), early metaphase (550 band stage) and elongated mid-metaphase (400 band stage), are routinely obtained. Technical improvements brought about by this high-resolution chromosomal technique have shown that most cases with adult acute non-lymphocytic leukemia have abnormal karyotypes (19 out of 20 = 95% of cases) and new leukemic subgroups with specific chromosomal defects can be found. Since nearly 50% of all cases of ANLL were previously considered to have a normal karyotype, the new findings make possible a more precise diagnosis for a better assessment of the clinical course and response to treatment of leukemic patients. Recently we have developed a G-banded late and mid-prophase technique to obtain 1200 to 2000 bands per haploid set. Since the human genome is believed to contain only some 30,000 genes, the application of this technique to the study of leukemias should provide an unprecedented level of sophistication in cancer cytogenetics. This technique, for example, would make it possible to visualize a deletion, duplication or inversion involving one or a few subbands and define the precise breakpoints involved in the specific reciprocal translocations found in various neoplasias, postulated to involve specific "cancer gene" sites.