Purpose: A considerable part of ductal carcinoma in situ (DCIS) lesions may never progress into invasive breast cancer. However, standard treatment consists of surgical excision. Trials aim to identify a subgroup of low-risk DCIS patients that can safely forgo surgical treatment based on histologic grade, which highlights the importance of accurate grading. Using real-life nationwide data, we aimed to create insight and awareness in grading variation of DCIS in daily clinical practice. Methods: All synoptic pathology reports of pure DCIS resection specimens between 2013 and 2016 were retrieved from PALGA, the nationwide Dutch Pathology Registry. Absolute differences in proportions of grade I-III were visualized using funnel plots. Multivariable analysis was performed by logistic regression to correct for case-mix, providing odds ratios and 95% confidence intervals for high-grade (III) versus low-grade (I–II) DCIS. Results: 4952 DCIS reports from 36 laboratories were included, of which 12.5% were reported as grade I (range 6.1–24.4%), 39.5% as grade II (18.2–57.6%), and 48.0% as grade III (30.2–72.7%). After correction for case-mix, 14 laboratories (38.9%) reported a significantly lower (n = 4) or higher (n = 10) proportion of high-grade DCIS than the reference laboratory. Adjusted ORs (95%CI) ranged from 0.52 (0.31–0.87) to 3.83 (1.42–10.39). Significant grading differences were also observed among pathologists within laboratories. Conclusion: In this cohort of 4901 patients, we observed substantial inter- and intra-laboratory variation in DCIS grading, not explained by differences in case-mix. Therefore, there is an urgent need for nationwide standardization of grading practices, especially since the future management of DCIS may alter significantly depending on histologic grade.