Photosynthesis and respiration impart distinct isotopic signatures to the atmosphere that are used to constrain global carbon source/sink estimates and partition ecosystem fluxes. Increasingly, the Keeling plot method is being used to determine the carbon isotope composition of ecosystem respiration (δ13CR) in order to better understand the processes controlling ecosystem isotope discrimination. In this paper we synthesize emergent patterns in δ13CR by analyzing 146 Keeling plots constructed at 33 sites across North and South America. In order to interpret results from disparate studies, we discuss the assumptions underlying the Keeling plot method and recommend standardized methods for determining δ13CR. These include the use of regression calculations that account for error in the x variable, and constraining estimates of δ13CR to nighttime periods. We then recalculate δ13CR uniformly for all sites. We found a high degree of temporal and spatial variability in C3 ecosystems, with individual observations ranging from -19.0 to -32.6?. Mean C3 ecosystem discrimination was 18.3?. Precipitation was a major driver of both temporal and spatial variability of δ13CR, suggesting (1) a large influence of recently fixed carbon on ecosystem respiration and (2) a significant effect of previous climatic effects on δ13CR. These results illustrate the importance of water availability as a key control on atmospheric 13CO2 and highlight the potential of δ13CR as a useful tool for integrating environmental effects on dynamic canopy and ecosystem processes.