Background/Objectives: The precise staging of non-small cell lung cancer (NSCLC) determines its initial treatment and provides more accurate prognostic information for patients. The aim of this cohort study was to determine pre- and post-operative mediastinal nodal staging agreement and its effect on 2-year survival. Methods: A retrospective multi-centre cohort study was performed, using prospectively collected and pre-defined data from weekly lung cancer multidisciplinary team (MDT) meetings in 11 hospitals. Consecutive patients who underwent surgical resection of NSCLC between 2015 and 2017 were eligible. Pre-operative under-staging was defined as a lower pre-operative than post-operative nodal stage, and pre-operative over-staging as a higher pre-operative than post-operative nodal stage. Disparities between pre-operative nodal staging established at MDT and post-surgical nodal staging were determined and associations with subsequent lung cancer-specific 2-year mortality were identified using univariate and multivariate regression. Results: A total of 973 patients fulfilled the eligibility criteria. Concordant pre- and post-operative nodal staging was observed in 783/973 (80%), 123/973 (13%) were under-staged pre-operatively and 67/973 (7%) were over-staged. In 173 patients with clinical N1 or N2 disease (in whom invasive mediastinal staging was indicated), staging EBUS was performed in 55/173 (32%). In these patients, younger age and use of staging EBUS were independent predictors of concordant pre- and post-operative staging. In all patients, pre-operative under-staging was independently associated with increased lung cancer-specific 2-year mortality. There was no increased mortality associated with pre-operative nodal over-staging. Conclusions: Invasive mediastinal staging with EBUS was independently associated with more accurate pre-operative staging. Pre-operative nodal under-staging was associated with increased lung cancer-specific mortality. Nodal staging accuracy in potentially curable NSCLC is of fundamental importance to ensure patients receive the correct first-line treatment and to improve survival.