In a 4-year period 168 tumor specimens received from 119 pediatric patients were plated in a soft agar cloning system. Overall, 47 or 28% of the tumor specimens had growth adequate enough for drug sensitivity testing. Drug sensitivity testing was performed using a variety of standards as well as investigational anticancer agents. Overall, 129 evaluable in vitro drug sensitivity tests were performed with 33 (26%) of the tests showing sensitivity to an agent. The most active drugs in vitro included the standard agents doxorubicin and cis-platinum as well as the investigational agents mitoxantrone and m-AMSA. From these data it is clear the cloning assay can be used to study pediatric malignancies. However, before the cloning assay can be applied to clinical pediatric oncology practice, the assay must be improved in terms of tumor growth and validated with prospective clinical trials.