BACKGROUND: Iron deficiency is widely underdiagnosed in female college athletes and may limit their athletic performance. Routine screening is necessary to identify potential female athletes who might benefit from iron supplementation. A retrospective study on the prevalence of iron deficiency in female collegiate division I athletes is imperative to bring awareness to the prevailing problem. The objective of this study is to determine the prevalence of iron deficiency in female athletes presenting to a Division 1 Athletic Training Room for initial health screening evaluation from January 2017 to July 2022. METHODS: A retrospective chart review was performed for female athletes who presented for initial pre-participation examination health screening to a Division 1 Athletic Training Room The data collected included the patients’ age, BMI, menstrual history, ferritin, and hemoglobin lab values. The primary sports analyzed was swim and dive, soccer, basketball, track and field/cross country, softball, lacrosse, volleyball, high tech, tennis, golf, and cheerleading. Descriptive statistics were used to describe the prevalence of iron deficiency among female athletes and the Student’s t-test and Pearson correlation analysis are used to identify any potential risk factors for iron deficiency. RESULTS: There was a total of 336 participants. The prevalence of athletes with low initial ferritin values under 40 ng/mL at pre-participation examination was 194 (57.74%, n=336). The mean ferritin value at pre-participation examination was 43.18 ng/mL (n=336). Statistical analysis of the other variables collected in this study including body mass index, menstrual history and primary sport showed no significant differences in relationship to initial ferritin levels. Female athletes with low ferritin levels were more likely to receive iron supplementation (p=8.089e-18, 95% CI= -31.33142, -20.19980). An increase in ferritin levels was correlated with iron supplementation (p=0.0217, 95% CI= 1.45825, 18.34014) and hemoglobin levels were higher in female athletes with higher ferritin levels (p-value=0.04971, 95% CI=0.00546, 0.14506). Initial ferritin levels were positively correlated with hemoglobin levels (p=2.402e-11, 95% CI=0.26962, 0.46399). Statistical analysis showed an increase in ferritin levels with usage of contraception (p=0.0073, 95% CI= 3.15448, 20.05362). CONCLUSION: This study’s findings represent the importance of screening ferritin levels in all female collegiate athletes regardless of sport, BMI, or menstrual history due to the high prevalence of low ferritin values. There was an increase in ferritin levels with usage of birth control suggesting the need to further evaluate the impact of menstrual history and contraception usage on ferritin and hemoglobin levels. In these authors experience, it is uncommon for institutions to incorporate routine screening measures in place for screening all female athletes with a ferritin value and complete blood count at initial pre-participation examination.