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Background: The clinical presentation of adnexal torsion in children is nonspecific. In some studies, serum D-dimer showed promise as a biochemical marker. Thus, this multi-center prospective observational study aimed to evaluate the sensitivity and specificity of preoperative serum D-dimer levels for diagnosing adnexal torsion in children and adolescents. Methods: This study prospectively enrolled female patients aged < 18 years presenting to the emergency departments of participating centers with symptoms suggestive of adnexal torsion between January 2022 and December 2024. Preoperative serum D-dimer levels were measured for all patients undergoing surgical exploration. Patients’ characteristics were examined using descriptive and inferential statistics, and the accuracy of preoperative serum D-dimer levels for diagnosing adnexal torsion was assessed using univariate logistic regression and a receiver operating characteristic curve. Results: This study enrolled 28 patients aged 4–17 years. Adnexal torsion was found in 17 patients, on the left side in 4 (23.53%) and on the right side in 13 (76.47%). Almost all patients were treated laparoscopically, and no postoperative complications occurred. Pre-operative serum D-dimer levels were higher among patients with adnexal torsion than among those without. The univariate model of serum D-dimer levels had an odds ratio of 1, a sensitivity of 0.77, and a specificity of 0.82 (p = 0.27). Conclusions: No direct association was observed between preoperative serum D-dimer levels and adnexal torsion. Nonetheless, the sensitivity and specificity suggest the possible utility of including serum D-dimer levels in multi-marker diagnostic models to complement rather than replace existing tools.