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Background: Hemoglobin A1c (HbA1c) levels play an important role in diagnosing, screening, and monitoring the treatment of diabetes. Our study aims to determine whether a relationship exists between HbA1c levels and age and gender in Turkish adults who have not been diagnosed with diabetes., Methods: This retrospective study included 6776 Turkish adults with no known diabetes. Cross-sectional analyses of A1C levels were performed between different age and gender categories. In statistical analysis, t-test, linear regression analysis, one-way ANOVA analysis, and LSD post hoc were used., Results: HbA1c levels in the individuals examined by dividing into different age groups increased with age in all groups. In our study, HbA1c levels were significantly higher in males than females (p<0.001) in all groups, especially between the ages of 30-49, and were positively associated with age for males and females. There was a positive correlation between HbA1c and age in both mans and women aged 30-49 (P<0.05). In the HbA1c 6.5% group newly diagnosed with diabetes, HbA1c levels gradually decreased with age in both genders, and no significant effect of age on HbA1c level was detected (p>0.05)., Conclusions: Our results showed that it is important to evaluate the effects of age and gender when using HbA1c levels in the diagnosis, screening, and treatment of diabetes, especially in young and middle-aged populations. Applying this situation to daily practice may reduce the misdiagnosis of diabetes in elderly patients, overtreatment of diabetes, and its associated risks., Competing Interests: All the authors declare that they have no conflict of interest in this work.Conflict of Interest: The authors stated that they have no conflicts of interest regarding the publication of this article., (2023 Şener Gülsen, Kahvecioğlu Esra Deniz, Can Başak, Gümüş Alper, Beyazıt Semih Yeşil, Evran Betül, published by CEON/CEES.)