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Objective: To find out the effect of iron deficiency anemia HbA1c levels in female patients with and without diabetes by comparing its values in groups. Study Design: Comparative cross-sectional study Place and Duration of Study: Physiology department, Al-Nafees Medical College and Pakistan Institute of Medical Sciences, Islamabad, from Jan 2017 to Dec 2018. Methodology: Study Participants were divided into four groups. Control group A, had females without diabetes and iron deficiency anemia whereas group B had anemic females without diabetes. Study subjects of groups C were having diabetes with anemia where group D females were with diabetes and without iron deficiency anemia. Values of HbA1c were compared in anemic and non-anemic groups, in females with and without diabetes by applying ANOVA and Post Hoc Tukey’s Test. Haemoglobin levels, HbA1c, erythrocyte indices and fasting blood glucose (FBG) were measured. Results: One hundred and forty adult females with and without diabetes were selected in the study. Significant p values were obtained when HbA1c levels of patients with anemia were compared with those without anemia in nondiabetics (p-value <0.00) as well as in diabetic group (p-value=0.048). Pearson correlation also showed significant negative correlation (r= -0.66, p-value <0.00) of haemoglobin levels with HbA1c. Conclusion: Iron deficiency anemia was found to be associated with an increase in HbA1c values in females with and without diabetes and hence can affect screening, diagnostic and prognostic efficacy of HbA1c for diabetes mellitus. [ABSTRACT FROM AUTHOR]