Tree-ring stable isotopes are typically measured in latewood cellulose to mitigate potential carry-over effects from previous year storage pools. The isotopic composition of individual tree-ring segments is thought to include considerable intra-annual variability. This sampling strategy may be complicated by steep intra-annual isotope gradients that can rival the inter-annual variability, however. Consistent sampling of latewood material may not always be possible due to low sample availability or high prevalence of narrow rings or low amounts of latewood because of species-specific changes in ring width. Therefore, years that contain samples with higher portions of non-latewood (earlywood) material may influence the final chronology of isotopic variability. Here, we analyze the potential influence that changing earlywood and latewood components of individual tree rings can have on stable carbon and oxygen records from Quercus spp. and Pinus heldreichii chronologies. Analysis of stable isotopes in oak tree rings with varying amounts of latewood show no statistically significant differences in the range of isotopic composition, nor any major differences when considering the same calendric year. Similar results were found for the pine data, when comparing stable isotope measurements with earlywood-to-latewood ratio and maximum density. We argue that this simple approach should be applied to any long-term tree-ring stable isotope record in order to provide a better understanding of the potential biases that could arise from previously recorded intra-annual variability in the wood.