Secondary forests (SFs) regenerating on previously deforested land account for large, expanding areas of tropical forest cover. Given that tropical forests rank among Earth’s most important reservoirs of carbon and biodiversity, SFs play an increasingly pivotal role in the carbon cycle and as potential habitat for forest biota. Nevertheless, their capacity to regain the biotic attributes of undisturbed primary forests (UPFs) remains poorly understood. Here, we provide a comprehensive assessment of SF recovery, using extensive tropical biodiversity, biomass, and environmental datasets. These data, collected in 59 naturally regenerating SFs and 30 co-located UPFs in the eastern Amazon, cover >1,600 large- and small-stemmed plant, bird, and dung beetles species and a suite of forest structure, landscape context, and topoedaphic predictors. After up to 40 years of regeneration, the SFs we surveyed showed a high degree of biodiversity resilience, recovering, on average among taxa, 88% and 85% mean UPF species richness and composition, respectively. Across the first 20 years of succession, the period for which we have accurate SF age data, biomass recovered at 1.2% per year, equivalent to a carbon uptake rate of 2.25 Mg/ha per year, while, on average, species richness and composition recovered at 2.6% and 2.3% per year, respectively. For all taxonomic groups, biomass was strongly associated with SF species distributions. However, other variables describing habitat complexity—canopy cover and understory stem density—were equally important occurrence predictors for most taxa. Species responses to biomass revealed a successional transition at approximately 75 Mg/ha, marking the influx of high-conservation-value forest species. Overall, our results show that naturally regenerating SFs can accumulate substantial amounts of carbon and support many forest species. However, given that the surveyed SFs failed to return to a typical UPF state, SFs are not substitutes for UPFs.