Monitoring crop growth and estimating crop yield are essential for managing agricultural production, ensuring food security, and maintaining sustainable agricultural development. Combining the mechanistic framework of a crop growth model with remote sensing observations can provide a means of generating realistic and spatially detailed crop growth information that can facilitate accurate crop yield estimates at different scales. The main objective of this study was to develop a robust estimation methodology of within-field winter wheat yield at a high spatial resolution (20 m × 20 m) by combining a light use efficiency-based model and Sentinel-2 data. For this purpose, Sentinel-2 derived leaf area index (LAI) time series were assimilated into the Simple Algorithm for Yield Estimation (SAFY) model using an ensemble Kalman filter (EnKF). The study was conducted on rainfed winter wheat fields in southern Sweden. LAI was estimated using vegetation indices (VIs) derived from Sentinel-2 data with semi-empirical models. The enhanced two-band vegetation index (EVI2) was found to be a useful VI for LAI estimation, with a coefficient of determination (R2) and a root mean square error (RMSE) of 0.80 and 0.65 m2/m2, respectively. Our findings demonstrate that the assimilation of LAI derived from Sentinel-2 into the SAFY model using EnKF enhances the estimation of within-field spatial variability of winter wheat yield by 70% compared to the baseline simulation without the assimilation of remotely sensed data. Additionally, the assimilation of LAI improves the accuracy of winter wheat yield estimation by decreasing the RMSE by 53%. This study demonstrates an approach towards practical applications of freely accessible Sentinel-2 data and a crop growth model through data assimilation for fine-scale mapping of crop yield. Such information is critical for quantifying the yield gap at the field scale, and to aid the optimization of management practices to increase crop production.