Abstract: Objectives To investigate the effects of anti-nuclear antibody (ANA) and nine anti-ENA antibodies (Ro-52, SSA, SSB, Jo-1, RNP/Sm, Sm, Scl-70, Sm, Scl-70, PM-Scl, ds DNA) on the detection of syphilis antibody in pregnant women. Methods A total of 253 cases was recruited, with 138 anti-nuclear antibody positive pregnant women (experimental group 1) , 55 anti-nuclear antibody negative pregnant women (experimental group 2) and 60 healthy non-pregnant women (healthy control group) and serum TP-Ab was detected with ELISA as screening method. The positive incidents of TP-Ab of ANA and its nine anti-ENA antibody were confirmed by TPPA method. Chi-square test was used to compared positive rate. Results There were 12 cases of TP-Ab positive in the experimental group 1, accounting for 8.70% (12/138) , which was significantly higher than that in the healthy control group (P<0.05) , while there was no significant difference (P>0.05) between experimental group 2 and control group, with TP-Ab rate of 1.82% (1/55) and zero re.positive antibody TP-Ab of Anti-ENV antibody were only detected in Ro-52 and SSA, which of positive incidence accounted for 14.71% (10/68) and 5.40% (2/37) respectively, there was significant difference between the two groups (P<0.05) . The results in 253 cases were totally confirmed negative through TPPA method. Conclusions ANA can interfere with the detection of TP-Ab, which caused false positive results, especially the risk of false positivity caused by Ro-52 is higher.