Background: To investigate the value of intraoperative frozen section examination (IFSE) in multiparametric magnetic resonance imaging/transrectal ultrasound (mpMRI/TRUS) fusion prostate biopsy in a major pandemic.Methods: A total of 35 patients were prospectively enrolled in our hospital from March 2020 to January 2021. The mpMRI/TRUS fusion system was used to perform a targeted biopsy, and the collected specimens were examined by IFSE (Observation Group 1). Then, a targeted biopsy was performed again for routine pathological examination (Observation Group 2). Finally, a systemic biopsy was performed, and the obtained specimens were routinely examined (Control Group). The positive rate, single core positive rate, Gleason score, and time to obtain pathological reports were compared between the groups.Results: The positive rate was 48.6% (17/35) in the control group, 48.6% (17/35) in Observation Group 1, and 51.4% (18/35) in Observation Group 2, showing no significant difference (P>0.05). The single core positive rates were 17.8%, 44.6%, and 47.1% in the Control Group, Observation Group 1, and Observation Group 2, respectively. Observation Group 1 and Observation Group 2 were significantly different from the Control Group (P<0.001). No participants in Observation Group 1 had increased or decreased Gleason scores compared with those in Observation Group 2. The time to obtain the pathological report was 0.025±0.014 days and 4.216±1.073 days for Observation Group 1 and Observation Group 2, respectively, showing a significant difference (P<0.001).Conclusions: This study showed that IFSE can not only rapidly obtain the pathological report of an mpMRI/TRUS biopsy, but can also ensure the accuracy of the pathological diagnosis.