We previously demonstrated that N-acetylcysteine (NAC) could reduce the toxicity of silver (Ag) materials (nanoparticles (NPs) and Ag nitrate) to the soil invertebrate Enchytraeus crypticus (Oligochaeta). It remains however, unclear whether the antitoxic mechanism of NAC was caused by NAC-Ag binding in the soil or inside the organisms. This study aimed at determining the bioavailability of Ag in the soil in a 21-day toxicity test as well as the Ag uptake and elimination kinetics in E. crypticus exposed to AgNPs in LUFA 2.2 standard soil amended with low (100 mg/kg dry soil) and high (600 mg/kg dry soil) NAC concentrations. The addition of NAC to the soil alleviated the toxicity of AgNPs by decreasing the internal Ag concentration of E. crypticus in a dose-dependent manner. Indeed, NAC reduced the binding of Ag to the soil, which probably was due to the formation of soluble but biologically unavailable Ag-cysteine complexes. The reduced Ag uptake in the enchytraeids was explained from an increased elimination at high NAC levels. These findings reinforce the view that metal complexing-compounds like NAC play a key role in the modulation of AgNP toxicity and bioavailability in terrestrial environments. Further, it may inform on the potential of NAC as a remediation solution for Ag or other metal-contaminated soils.