Background: Pulmonary embolism (PE) is a common disease associated with high mortality and morbidity. Diagnosing PE is challenging due to diverse clinical presentations and the lack of specific biomarkers. Objective: We hypothesized that plasma galectin-3 (Gal-3) levels might reflect the severity of acute PE and be useful for diagnostic assessment. Methods: In this prospective study, 150 patients (100 patients with PE and 50 control patients) were included. Patients were stratified into high-risk, medium-risk, and low-risk groups according to the Wells and revised Geneva scoring systems, and Gal-3 levels were compared among the groups. PE was diagnosed by means of computed tomography pulmonary angiography. Results: In this study, of the 100 PE patients included in the study, 69 patients recovered and were discharged and 31 patients died. Median Gal-3 value in the PE group was 27.0 ng/mL (range 11.5–35.0 ng/mL), whereas the median Gal-3 value in the control group was significantly lower at 8.8 ng/mL (range 1.0–21.0 ng/mL) (p < 0.001). When the Gal-3 values of the PE group and the control group were evaluated with the receiver operator characteristic curve, the area under the curve was calculated as 0.99 (95% confidence interval 0.979–1). At a Gal-3 cutoff value of 13.55 ng/mL, which was determined to be the most appropriate value for PE diagnosis, the sensitivity was 98% and the specificity was 92%. Conclusions: A biomarker that rapidly and accurately diagnoses acute PE in the emergency department can be an extremely useful tool. We concluded that plasma Gal-3 levels can be regarded as a promising marker of acute PE.