Objective To evaluate the effectiveness of register based, yearly chlamydia screening. Design Controlled trial with randomised stepped wedge implementation in three blocks. Setting Three regions of the Netherlands: Amsterdam, Rotterdam, and South Limburg. Participants 317 304 women and men aged 16-29 years listed on municipal registers at start of trial. Intervention From March 2008 to February 2011, the Chlamydia Screening Implementation programme offered yearly chlamydia screening tests. Postal invitations asked people to use an internet site to request a kit for self collection of samples, which would then be sent to regional laboratories for testing. Treatment and partner notification were done by the general practitioner or at a sexually transmitted infection clinic. Main outcome measures Primary outcomes were the percentage of chlamydia tests positive (positivity), percentage of invitees returning a specimen (uptake), and estimated chlamydia prevalence. Secondary outcomes were positivity according to sex, age, region, and sociodemographic factors; adherence to screening invitations; and incidence of self reported pelvic inflammatory disease. Results The participation rate was 16.1% (43 358/269 273) after the first invitation, 10.8% after the second, and 9.5% after the third, compared with 13.0% (6223/48 031) in the control block invited at the end of round two of the intervention. Chlamydia positivity in the intervention blocks at the first invitation was the same as in the control block (4.3%) and 0.2% lower at the third invitation (odds ratio 0.96 (95% confidence interval 0.83 to 1.10)). No substantial decreases in positivity were seen after three screening rounds in any region or sociodemographic group. Among the people who participated three times (2.8% of all invitees), positivity fell from 5.9% to 2.9% (odds ratio 0.49 (0.47 to 0.50)). Conclusions There was no statistical evidence of an impact on chlamydia positivity rates or estimated population prevalence from the Chlamydia Screening Implementation programme after three years at the participation levels obtained. The current evidence does not support a national roll out of this register based chlamydia screening programme. Trial registration NTR 3071 (Netherlands Trial Register, www.trialregister.nl).