Background: Pertussis is a highly infectious respiratory illness caused by the bacteria Bordetella pertussis. A resurgence of pertussis, even in countries with good vaccine coverage, has led to an increase in infant deaths. In response to this, many countries have introduced pertussis vaccination in pregnancy. This strategy is effective at preventing infant disease, but there remains uncertainty about what gestational timing is best to ensure maximal protection of the infant. These uncertainties are the rationale for this randomised controlled trial and a sub-study investigating pertussis-specific antibody in breastmilk. Protocol: We will recruit 354 pregnant women and will randomise them to receive their pertussis vaccination in one of three gestational age windows: ≤23+6, 24-27+6 and 28-31+6 weeks of gestation. Vaccination will be with Boostrix-IPV® and participants will be asked to complete a symptom diary for seven days following vaccination. Blood sampling will be performed prior to vaccination, two weeks following vaccination and at the time of delivery. A cord blood sample will be collected at delivery and a blood sample collected from the infant 4-10 weeks after completion of the primary immunisations. Individuals participating in the breastmilk sub-study will provide a sample of colostrum within 48 hours of delivery and samples of breastmilk at two weeks and around five-six months. Blood samples will be analysed using enzyme linked immunosorbent assay (ELISA) techniques for pertussis toxin, filamentous haemagglutinin and pertactin. A subset of serum samples will also be analysed using a functional assay. Colostrum and breastmilk samples will be analysed using functional assays. Discussion: Although pertussis vaccination has been shown to be safe and effective in pregnancy there remains debate about the optimal timing for the administration during pregnancy. This study will investigate antibody responses in serum and breastmilk when vaccination is performed in three different time periods. Clinicaltrials.gov registration: NCT03908164 (09/04/2019)