Stem biomass is a fundamental component of the global carbon cycle that is essential for forest productivity estimation. Over the last few decades, Light Detection and Ranging (LiDAR) has proven to be a useful tool for accurate carbon stock and biomass estimation in various biomes. The aim of this study was to investigate the potential of multispectral LiDAR data for the reliable estimation of single-tree total and barkless stem biomass (TSB and BSB) in an uneven-aged structured forest with complex topography. Destructive and non-destructive field measurements were collected for a total of 67 dominant and co-dominant Abies borisii-regis trees located in a mountainous area in Greece. Subsequently, two allometric equations were constructed to enrich the reference data with non-destructively sampled trees. Five different regression algorithms were tested for single-tree BSB and TSB estimation using height (height percentiles and bicentiles, max and average height) and intensity (skewness, standard deviation and average intensity) LiDAR-derived metrics: Generalized Linear Models (GLMs), Gaussian Process (GP), Random Forest (RF), Support Vector Regression (SVR) and Extreme Gradient Boosting (XGBoost). The results showcased that the RF algorithm provided the best overall predictive performance in both BSB (i.e., RMSE = 175.76 kg and R2 = 0.78) and TSB (i.e., RMSE = 211.16 kg and R2 = 0.65) cases. Our work demonstrates that BSB can be estimated with moderate to high accuracy using all the tested algorithms, contrary to the TSB, where only three algorithms (RF, SVR and GP) can adequately provide accurate TSB predictions due to bark irregularities along the stems. Overall, the multispectral LiDAR data provide accurate stem biomass estimates, the general applicability of which should be further tested in different biomes and ecosystems.