OBJECTIVE: Traditional non-invasive prenatal testing (NIPT) for sex chromosome aneuploidies (SCAs) has a higher risk of false positives, particularly for 45,X. This study introduces an innovative NIPT analysis pipeline to minimize false positives in 45,X screening. METHODS: We included 39 cases from the NIPT records of 15,882 pregnancies, including 20 with 45,X and 19 controls, and performed standard NIPT and size-selection NIPT tests. Based on the two NIPT results, the Z RESULTS: By integrating data from two NIPT experiments and monitoring sex chromosome variations, the Z CONCLUSION: Our advanced analysis process significantly improved the PPV of NIPT for 45,X and reduced the incidence of false positives.