BACKGROUND & AIMS The fecal immunochemical test (FIT) is superior to the guaiac-based fecal occult blood test in detecting neoplasia. There are not much data on the optimal number of FITs to perform. We conducted a population-based trial to determine attendance and diagnostic yield of 1- and 2-sample FIT screening. METHODS The study included 2 randomly selected groups of subjects aged 50-74 years (1-sample FIT, n=5007; 2-sample FIT, n=3197). The 2-sample group was instructed to collect fecal samples on 2 consecutive days. Subjects were referred for colonoscopy when at least 1 sample tested positive (≥50 ng hemoglobin/mL). RESULTS Attendance was 61.5% in the 1-sample group (2979 of 4845; 95% confidence interval, 60.1%-62.9%) and 61.3% in the 2-sample group (1875 of 3061; 95% confidence interval, 59.6%-63.0%; P=.84). In the 1-sample group 8.1% tested positive, and in the 2-sample group 12.8% had at least 1 positive test outcome and 5.0% had 2 positive test outcomes (P<.05). When the mean from both test results in the 2-sample group was used, 10.1% had a positive test outcome (P<.05). The detection rates for advanced neoplasia were 3.1% in the 1-sample group, 4.1% in the 2-sample group with at least 1 positive test outcome, 2.5% when both test results were positive, and 3.7% among subjects with the mean from both test results being positive. CONCLUSIONS There is no difference in attendance for subjects offered 1- or 2-sample FIT screening. The results allow for the development of efficient FIT screening strategies that can be adapted for local colonoscopy capacities, rather than varying the cut-off value in a 1-sample strategy.