Background The prothrombotic defect factor V Leiden (FVL) may confer higher risk of ST‐segment–elevation myocardial infarction (STEMI), compared with non–ST‐segment–elevation acute coronary syndrome, and may be associated with more myocardial necrosis caused by higher thrombotic burden. Methods and Results Patients without history of cardiovascular disease were selected from 2 clinical trials conducted in patients with acute coronary syndrome. FVL was defined as G‐to‐A substitution at nucleotide 1691 in the factor V (factor V R506Q) gene. Odds ratios were calculated for the association of FVL with STEMI adjusted for age and sex in the overall population and in the subgroups including sex, age (≥70 versus <70 years), and traditional cardiovascular risk factors. The peak biomarker levels (ie, creatine kinase‐myocardial band and high‐sensitivity troponin I or T) after STEMI were contrasted between FVL carriers and noncarriers. Because of differences in troponin assays, peak high‐sensitivity troponin levels were converted to a ratio scale. The prevalence of FVL mutation was comparable in patients with STEMI (6.0%) and non–ST‐segment–elevation acute coronary syndrome (5.8%). The corresponding sex‐ and age‐adjusted odds ratio was 1.06 (95% CI, 0.86–1.30; P =0.59) for the association of FVL with STEMI. Subgroup analysis did not show any differences. In patients with STEMI, neither the median peak creatine kinase‐myocardial band nor the peak high‐sensitivity troponin ratio showed any differences between wild‐type and FVL carriers ( P for difference: creatine kinase‐myocardial band=0.33; high sensitivity troponin ratio=0.54). Conclusions In a general population with acute coronary syndrome, FVL did not discriminate between a STEMI or non–ST‐segment–elevation acute coronary syndrome presentation and was unrelated to peak cardiac necrosis markers in patients with STEMI. Registration URL: https://www.clinicaltrials.gov ; Unique identifiers: NCT00391872 and NCT01761786.