OBJECTIVES: The rise in molecular diagnostic testing in oncology has made these tests the subject of an increasing number of investment discussions, reimbursement negotiations and hospital cost calculations. This requires a detailed assessment of the total cost for determining tumor mutation status beyond plain kit costs. This research maps the workflow, identifies investment needs and quantifies the direct variable costs of KRAS and NRASmutation testing from formalin fixed paraffin embedded (FFPE) sample to result. METHODS: The complete test workflow is based on the current workflow in two academic European institutions. Hands-on time and consumables used in each step were quantified in both hospitals. Unit cost was obtained from the finance departments. RESULTS: Costs related to tumor mutation tests include investments (lab space, lab equipment, molecular diagnostic systems, training, quality assurance, accreditation), indirect costs, lab running costs (disinfection, cost of errors and repeats) and direct variable costs including reagent costs, controls and labor. The total cost depends on workflow and tumor profile. To detect a wild-type RAS tumor, the direct variable costs from FFPE sample to result excluding reagent costs, is estimated at €64 using a CE-IVD kit for KRAS exon 2 followed by pyrosequencing for NRAS exon 2 and extended RAS testing, €51 using High Resolution Melting confirmed by pyrosequencing and €3 for an automated instrument. The latter automated technology includes sample preparation in the test cost, as it works directly from FFPE. CONCLUSIONS: Estimations on the cost of performing tumor mutation tests should include not only reagent costs, which are usually considered, but also investments, lab running costs, labor and all other direct and indirect costs. Even in the most advanced labs, efforts are needed to fully understand the total cost of ownership as this will become ever more important for future market access, reimbursement and financial discussions.