Abstract Background Children and adolescents spend a lot of time sitting at school. Implementing standing desks in the classroom is one potential strategy to reduce and break up sitting time. The first aim was to evaluate the effect of implementing standing desks in classrooms in primary and secondary schools on pupils’ sitting-related behaviour and determinants. The second aim was to quantitatively and qualitatively evaluate the process of implementing the desks in the classroom. Methods We conducted a cluster-randomised controlled trial with a pre-, mid-, and post-test design including 10 intervention schools (5 primary, 5 secondary schools) and 9 control schools (5 primary, 4 secondary schools) across Flanders, Belgium. Three standing desks were placed in one class in each intervention school for 6 months. At pre-, mid- and post-test, all pupils (n = 311; 54.5% girls) completed a questionnaire whilst a subsample of three pupils per class wore an activPAL inclinometer for one school week. Focus groups with pupils and interviews with teachers were conducted at mid-test. Process evaluation questions were added to the mid- and post-test questionnaire for the intervention group. Qualitative data were analysed using NVivo 11. Multilevel regression analyses were conducted in MLwiN 2.31. Results Few significant intervention effects were observed, although activPAL data showed favourable intervention effects on primary school pupils’ sitting and standing time and bouts. Focus groups and interviews showed a generally positive attitude towards using standing desks in both teachers and pupils, although some barriers and suggestions for future implementation were noted, for example regarding the amount of desks per classroom. Quantitative process evaluation data showed a low individual use of standing desks (between 57 and 83 min per week), which significantly decreased across the school year for primary school pupils only. Conclusions Although pupils and teachers were generally positive about the desks, relatively few ...