The physiological and biomechanical changes that occur during pregnancy make accurate measurement of physical activity (PA) a challenge during this unique period. The Global Physical Activity Questionnaire (GPAQ) has been used extensively in low-to-middle income countries, but has never been validated in a pregnant population. In this longitudinal study, 95 pregnant women (mean age: 29.5±5.7 years; BMI: 26.9±5.0 kg/m2) completed the GPAQ and were asked to wear an accelerometer for 7 days at two time points during pregnancy (14-18 and 29-33 weeks gestation). There was a significant difference between accelerometry and GPAQ when measuring moderate-to-vigorous physical activity (MVPA) at 29-33 weeks gestation (16.6 vs 21.4 min/day; p = 0.02) as well as sedentary behaviour (SB) at both 14-18 weeks (457.0 vs 300 min/day; p < 0.01) and 29-33 weeks gestation (431.5 vs 300 min/day; p < 0.01). There was poor agreement between the GPAQ and accelerometry for both PA and SB at both time points (ICC: -0.05-0.08). Bland Altman plots indicated that the GPAQ overestimates PA by 14.8 min/day at 14-18 weeks and by 15.8 min/day at 29-33 weeks gestation. It underestimates SB by 127.5 min/day at 14-18 weeks and by 89.2 min/day at 29-33 weeks gestation. When compared to accelerometry, the GPAQ shows poor agreement and appears to overestimate PA and underestimate SB during pregnancy.