The growing interest in the molecular subclassification of colorectal cancers is increasingly facilitated by large multicenter biobanking initiatives. The quality of tissue sampling is pivotal for successful translational research. This study shows the quality of fresh frozen tissue sampling within a multicenter cohort study for colorectal cancer (CRC) patients. Each of the seven participating hospitals randomly contributed ten tissue samples, which were collected following Standard Operating Procedures (SOP) using established techniques. To indicate if the amount of intact RNA is sufficient for molecular discovery research and prove SOP compliance, the RNA integrity number (RIN) was determined. Samples with a RIN < 6 were measured a second time and when consistently low a third time. The highest RIN was used for further analysis. 91% of the tissue samples had a RIN ≥ 6 (91%). The remaining six samples had a RIN between 5 and 6 (4.5%) or lower than 5 (4.5%). The median overall RIN was 7.3 (range 2.9–9.0). The median RIN of samples in the university hospital homing the biobank was 7.7 and the median RIN for the teaching hospitals was 7.3, ranging from 6.5 to 7.8. No differences were found in the outcome of different hospitals (p = 0.39). This study shows that the collection of high quality fresh frozen samples of colorectal cancers is feasible in a multicenter design with complete SOP adherence. Thus, using basic sampling techniques large patient cohorts can be organized for predictive and prognostic (bio)marker research for CRC.