Flow cytometric analysis of peripheral blood and bone marrow for tumor cells in patients with neuroblastoma

Abstract

Several sensitive surveillance tests reportedly have been used to detect occult neuroblastoma (NB) cells in peripheral blood (PB) and bone marrow (BM). They may be useful in monitoring minimal residual tumor cells. The authors report the feasibility and clinical usefulness of a sensitive flow cytometric assay that has been newly developed and evaluated to detect NB cells. Nine NB patients and 15 healthy donors were included in the current study. Primary tumor tissues, BM, and PB were examined for the detection of NB cells using a triple-color flow cytometric assay. Tumor cells in PB and BM, isolated by fluorescence-activated cell sorting, were used for morphologic studies and differential polymerase chain reaction analysis of N-myc gene amplification. Neuroblastoma cells consistently showed CD9+/CD56+/CD45- phenotype. Flow cytometric analysis could detect NB cells at a level of 1 per 10(4-5) cells. The CD9+/CD56+/CD45- cell population was absent in normal PB and BM. This assay identified occult NB cells, which were not detected by conventional cytology, in PB and BM obtained from six patients (one of two with Stage II and all five with advanced disease) at diagnosis. Residual NB cells also were detected in PB and BM during therapy. Neuroblast-like morphology and N-myc gene amplification of sorted cells confirmed that CD9+/CD56+/CD45- cells were truly NB cells. A triple-color flow cytometric assay was a sensitive and specific method to detect occult NB cells in PB and BM. This assay could be an additional component of surveillance testing for NB patients.