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Sedentary time questionnaires provide a low cost, low participant burden way of assessing sedentary behavior, but their subjectivity is questionable. The aim of this study was to assess the validity of sedentary time questionnaires, both single item and domain specific, in a student population under free living conditions with accelerometry as criterion measure. It is hypothesised that both sedentary time questionnaires underestimate the sitting time. Twenty healthy subjects (15 male, 5 female) participated in a one-week observational study under free-living conditions. They wore an accelerometer (activPAL) continuously for 7 consecutive days and afterwards filled in a single item (IPAQ) and a domain specific sedentary time questionnaire (dsSTQ) about the same time period. Spearman correlations were used to assess relative validity and Wilcoxon signed-rank tests and Bland-Altman plots were used to assess absolute agreement between questionnaires and accelerometry. Sitting time was significantly underestimated by the IPAQ and insignificantly overestimated by the dsSTQ compared to accelerometry for the full week average sitting times. Correlations between questionnaires and accelerometry were weak to moderate. For both questionnaires there were large individual differences in estimations compared to accelerometry. The correlation between questionnaire and accelerometer determined sitting events lasting 30 minutes or longer was moderate. The dsSTQ is the more accurate measure of the two questionnaires on sitting time on the population level. However, there were large individual differences in estimations, and thus it does not seem to be a valid, reliable measurement tool for sitting time in small populations and in studies were sitting time is an important outcome. For these studies, accelerometry or more preferably direct observation provides a better estimate of sitting time. The validity of single item and domain specific sitting time questionnaires in a student population measured under free-living conditions T.D. Atsma MaRBLe Research Papers 18