OBJECTIVE: Non-invasive prenatal testing (NIPT) can determine fetal chromosomal sex early in pregnancy, raising the possibility of sex selection. However, current evidence regarding this practice is limited. Our objective was to assess the sex ratio at birth (SRB) among a cohort of infants born following NIPT from a single laboratory provider in Victoria, Australia (2014-2018). METHOD: The NIPT dataset was linked with Victorian birth records. SRB and estimated relative risk of the birth of a male infant based on maternal major region of birth, parity and maternal age were calculated. An SRB was considered outside the natural range if the point estimate and 95% confidence interval were outside 1.04-1.06. RESULTS: The final dataset for analysis contained n = 43,665 NIPT records. The majority (88.5%, n = 38,631) were linked with a birth record. The overall SRB was within the natural range (1.05, 95% CI 1.029-1.071). Confidence intervals for all maternal major regions of birth, parities, and maternal age categories overlapped with the natural range. There was an increased estimated relative risk of a male birth for mothers born in Southern and Central Asia compared with mothers born in Australia (RR: 1.049, 95% CI 1.005-1.096, p = 0.03). CONCLUSION: We did not find evidence of an overall skewed SRB among a cohort of Australian NIPT users. However, our findings warrant larger studies on NIPT use and possible sex selection practices.