OBJECTIVES: The aim of this study is to determine the frequency of alleles and genotypes for mutations C282Y, H63D, S65C, HFE genotypes, and differences in the frequency of alleles and genotypes with regard to the sex of the subjects. STUDY DESIGN: Cross-sectional study. SUBJECTS AND METHODS: The studied group consists of 100 patients suspected of HH. Patients underwent molecular diagnostics in the Laboratory for Molecular and HLA Diagnostics of the Clinical Hospital Center in Osijek in the period from 2017 to 2021. Genomic DNA was isolated from whole blood samples with anticoagulant, and DNA concentration and purity were measured. Genotyping of the C282Y, H63D and S65C mutations in the HFE gene was performed using real-time polymerase chain reaction and melting curve analysis. Data on HFE genotyping results were collected by searching the e-Delphyn electronic database. RESULTS: In the group of subjects for the C282Y mutation, the most represented genotype is GG (74.0%) and allele G (83.0%), for the H63D mutation, the most represented genotype is CC (68.0%) and allele C (82.5%), and for the S65C mutation, the most common genotype is AA (97.0%) and allele A (98.5%). In the studied group of patients, there are 8 genotypes due to mutations C282Y, H63D and S65C in the HFE gene. The highest frequency is the genotype without HFE mutations (46.0%). CONCLUSION: In the age category between 51 and 70 years of age, a significantly larger number of women were referred for molecular diagnosis of HFE. There is no statistically significant difference in the distribution of alleles and genotypes for mutations C282Y, H63D and S65C, nor in the distribution of HFE genotypes between male and female gender groups.