Ecological subsidies have been hypothesized to be key factors affecting the dynamics of the recipient ecosystems. We assessed the impact of the flood regime on the assimilation of a trophic subsidy produced in the floodplain of a large fluvial lake, Lake Saint-Pierre (Quebec, Canada) on five invertivore littoral fish species (yellow perch (Perca flavescens), pumpkinseed (Lepomis gibbosus), bridle shiner (Notropis bifrenatus), golden shiner (Notemigonus crysoleucas), and banded killifish (Fundulus diaphanus)). We hypothesized that the incorporation of floodplain-derived carbon into fish biomass and individual fitness would increase in years with higher magnitude and longer duration floods. Studying four consecutive years with contrasting spring-flood events, we used a stable isotope mixing model to estimate floodplain contribution, and linear mixed models to analyze the impact of hydrology on floodplain contribution and fish body condition. Floodplain and main river’s invertebrates exhibited statistically distinct isotopic ratios (t test: p value < 0.001). Floodplain contribution and body condition were positively correlated with flooding intensity and duration for all species. Body condition was positively correlated with the assimilation of floodplain-subsidy for golden shiner and banded killifish. Our study identified flooding intensity as a main driver influencing the assimilation of floodplain-derived carbon by littoral fish and their fitness in a large river ecosystem.