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596 FLOW MICROFLUOROMETRY ANALYSIS OF DNA IN HUMAN NEUROBLASTOMA CELLS

More effective treatment for neuroblastoma is needed, and new therapies are being developed and tested. A rapid method for measuring cell cycle distribution of tumor cells might provide valuable information for selection and timing of therapeutic agents. Therefore, we developed flow microfluorometry (FMF) for measuring DNA content of neuroblastoma cells. Seven established human neuroblastoma cell lines were utilized. Neuroblastoma cells frequently grow in clumps, and these were dispersed with sonication. DNA content was determined after fixation of cells in ethanol, treatment with RNAse, and staining with propidium iodide (PI). The percentage of cells in G1, S, and G2+M was determined by computerized curve fitting analysis of the DNA distribution. For example, LA-N-1 cells in the logarhythmic growth phase consisted of 64% G1, 25.6% S, and 10.4 G2+M cells. FMF analysis of DNA in tumor cells growing in vivo can be complicated by normal cells present in the tumor. Therefore, we developed a tumor cell identification system using rabbit antiserum against neuroblastoma cell surface antigens and fluoresceinated goat anti-rabbit immunoglobulin. Neuroblastoma cells treated with these reagents, and then ethanol and PI were successfully analyzed for DNA content with FMF. This new method should allow analysis of neuroblastoma cells in primary tumors and various metastatic sites and may provide new information of therapeutic value.