OBJECTIVES: Beyond understanding the pure cost of genomic sequencing, the real costs associated with implementing next-generation sequencing (NGS) into clinical practice are currently unknown. To elucidate the real costs and to provide a potential benchmark for reimbursement, a pilot study was conducted to identify and valuate the resources related to conducting clinical cancer whole-exome (cWES), transcriptome, and targeted panel sequencing in a cohort of pediatric cancer patients. METHODS: A cost model was calculated using 25 pediatric cancer patients who underwent clinical genomic sequencing at Columbia University Medical Center. Our institutional workflow developed by the Precision in Pediatric Sequencing (PIPseq) Program in the Division of Pediatric Oncology and the Personalized Genomics Medicine Laboratory in the Department of Pathology guided the identification of resources and costs associated with NGS. RESULTS: CONCLUSIONS: Since the reimbursement landscape for clinical genomic sequencing is currently unknown, a comprehensive cost calculation reflecting resource utilization across the whole sequencing workflow including costs associated with directed therapy based on molecular profiling results is necessary. These data serve as a starting point toward identifying and valuating resources associated with NGS and serve as a first step toward demystifying reimbursement for clinical genomic sequencing in Pediatric Oncology.