The morphologic, immunologic and cytogenetic features of leukemic cells obtained at the time of first bone marrow relapse were compared with those obtained at initial diagnosis in 287 children with acute lymphoblastic leukemia (ALL) who were entered consecutively in a laboratory classification study of the Pediatric Oncology Group (POG). L1 to L2 shifts in French-American-British morphologic subtype were more common than the reverse (81/178 versus 15/61, p less than 0.001). A small but marginally significant number of cases acquired cytoplasmic granules at relapse, and 50 cases underwent a shift in periodic acid-Schiff reactivity that slightly favoured positive to negative. Shifts in immunophenotype were relatively rare, although shifts in cases with a pre-B phenotype to early pre-B ALL or vice versa occurred in about a third of pre-B cases. Loss of HLA-DR or the common ALL antigen occurred in 20 and 11% of cases, respectively. Of the 116 cases with analyzable karyotypes at diagnosis and relapse, 36 (31%) showed a change in karyotypes at relapse, usually from normal to pseudodiploid or hyperdiploid. Cytogenetic evidence for the emergence of a new clone after initial diagnosis was found in only one case. Analysis of the correlation of clinical and lymphoblast biologic features with event-free survival after an initial marrow relapse failed to demonstrate any prognostic significance for the changes identified in this study. T-cell immunophenotype proved to be the only factor significantly related to the outcome of retrieval therapy.