Background/aim: Medullary thyroid cancer (MTC) originates from parafollicular cells (C cell) and produces calcitonin (CT). Basal serum CT was used in the diagnosis and treatment of MTC. If basal CT level is 100 pg/mL or higher, it is likely to have MTC, but if basal CT level is below 10 pg/mL, the probability of developing thyroid disease is low. In cases with basal CT level between 10?100 pg/mL, pentagastrin-stimulated (PS) CT level is studied to evaluate MTC and C cell hyperplasia (CHH). This study aimed to determine cut-off value for basal and PS peak CT level for diagnosis of MTC. Materials and methods: We retrospectively reviewed files of patients presented to endocrine outpatient clinic of Ege University, Medicine School, between 2010 and 2019; 176 patients with basal CT level of 10?100 pg/mL and patients with PS test were included to the study. Results: The receiver operating characteristic curve (ROC) analysis was used to determine cut-off value for basal CT that can discriminate cases with MTC and those with nodular goiter. Cut-off value for basal CT was calculated as 46.5 pg/mL (specificity; 100 %, sensitivity; 74 %). In the ROC analysis for peak PS CT, cut-off value was calculated as 285 pg/mL (specificity:100 %; sensitivity:82 %). When peak CT level was > 290 pg/mL in PS test, both specificity and sensitivity for MTC were determined as 100 %. The PS peak CT level > 285 pg/ mL was significant for MTC diagnosis while range of 117?274 pg/mL was significant for CHH. Conclusion: In this study, cut-off value was calculated as 46.5 pg/mL for basal CT, whereas 285 pg/mL for PS peak CT in the diagnosis of preoperative MTC.