Flow cytometry and fluorescence in situ hybridization to detect residual neuroblastoma cells in bone marrow

Background. In patients with neuroblastoma morphological assessment of BM for residual NB cells is not precise, particularly when the number of tumor cells is small. Procedure. To develop a sensitive and rapid method of detecting NB cells in BM, we assessed the efficiency of flow cytometry (FCM) using markers CD9, CD56, and CD45. The percent of CD9+/CD56+/CD45- (NB phenotype) cells was determined by FCM in 41 samples (16 patients) at various time points. For confirmation fluorescence in situ hybridization (FISH) for 17q gain was performed. Results. Nineteen of the 22 (86%) samples that were negative by morphology were positive by FCM (> 0.006% CD9+/CD56+/CD45- cells). The longest time to complete the FCM study was 3 hr. In six FISH experiments the sorted CD9+/CD56+/CD45- Population had a higher percentage of cells with 17q gain (11.5-95%) compared to a CD56-/CD45+ internal control population (2-8%). Conclusions. Our preliminary results suggest that FCM determination of the percent of CD9+/CD56+/CD45- cells is an effective method of rapidly detecting NB cells in BM.