The present investigation evaluates the accuracy of estimating above-ground biomass (AGB) by means of two different sensors installed onboard an unmanned aerial vehicle (UAV) platform (DJI Inspire I) because the high costs of very high-resolution imagery provided by satellites or light detection and ranging (LiDAR) sensors often impede AGB estimation and the determination of other vegetation parameters. The sensors utilized included an RGB camera (ZENMUSE X3) and a multispectral camera (Parrot Sequoia), whose images were used for AGB estimation in a natural tropical mountain forest (TMF) in Southern Ecuador. The total area covered by the sensors included 80 ha at lower elevations characterized by a fast-changing topography and different vegetation covers. From the total area, a core study site of 24 ha was selected for AGB calculation, applying two different methods. The first method used the RGB images and applied the structure for motion (SfM) process to generate point clouds for a subsequent individual tree classification. Per the classification at tree level, tree height (H) and diameter at breast height (DBH) could be determined, which are necessary input parameters to calculate AGB (Mg ha−1) by means of a specific allometric equation for wet forests. The second method used the multispectral images to calculate the normalized difference vegetation index (NDVI), which is the basis for AGB estimation applying an equation for tropical evergreen forests. The obtained results were validated against a previous AGB estimation for the same area using LiDAR data. The study found two major results: (i) The NDVI-based AGB estimates obtained by multispectral drone imagery were less accurate due to the saturation effect in dense tropical forests, (ii) the photogrammetric approach using RGB images provided reliable AGB estimates comparable to expensive LiDAR surveys (R2: 0.85). However, the latter is only possible if an auxiliary digital terrain model (DTM) in very high resolution is available because in dense natural forests the terrain surface (DTM) is hardly detectable by passive sensors due to the canopy layer, which impedes ground detection.