Objective To analyze the expression levels and clinical significance of serum anti-RO52, Jo-1, Mi-2, PL-7 and PL-12 antibodies in polymyositis (PM). Methods A retrospective study was conducted on 60 PM patients admitted to the Affiliated Hospital of Guizhou Medical University from January 2015 to January 2017, and they were selected as the observation group, and another 60 healthy patients admitted to our outpatient physical examination center during the same period were selected as the control group. The positive rates of serum anti-RO52, Jo-1, Mi-2, PL-7 and PL-12 were detected and compared between the two groups. The positive rates of serum anti-RO52, Jo-1, Mi-2, PL-7 and PL-12 antibodies in the ILD group and the heart injury group were compared. 60 cases of PM patients were followed up for 5 years, and survival analysis was performed by Kaplan-Meire method. The survival curve was plotted using Graph Pad Prism8.0, and the area under the curve (AUC) was calculated by receiver operating curve (ROC). The predictive value of anti-RO52, Jo-1, Mi-2, PL-7 and PL-12 antibodies on the prognosis of PM patients was analyzed. Results The positive rates of anti-RO52, Jo-1, Mi-2, PL-7 and PL-12 antibodies in the observation group were higher than those in the control group (P＜0.05). The positive rates of serum anti-RO52, Jo-1, Mi-2, PL-7 and PL-12 antibodies in ILD group were higher than those in non-ILD group (P＜0.05). The positive rates of serum anti-RO52, Jo-1, Mi-2, PL-7 and PL-12 antibodies in patients with heart damage were higher than those in patients without heart damage (P＜0.05). The median survival time of 60 PM patients was 48.14 months, and the 5-year survival rate was 85.00%. Comparison of survival curves between negative and positive groups of anti-RO52, Jo-1, Mi-2, PL-7 and PL-12 antibodies (P＞0.05). The AUC of combined detection of anti-RO52, Jo-1, Mi-2, PL-7 and PL-12 antibodies to predict the prognosis of PM patients was 0.795 (95% CI: 0.711-0.963), The sensitivity (90.34%) and specificity (88.14%) of the combined detection of anti-RO52, Jo-1, Mi-2, PL-7 and PL-12 antibodies were higher than those of single detection (P＜0.05).Conclusion The positive rate of anti-RO52, Jo-1, Mi-2, PL-7 and PL-12 antibodies in PM patients is high. The combined detection of the above antibodies can improve the prognostic efficacy of PM patients and has important guiding value