The translation of the original seawater signal (i.e. ambient temperature and δ18Osw) into distinct chambers of a single shell of a foraminifer during calcification can influence our interpretation of surface ocean conditions of the past, when based upon oxygen and carbon stable isotope geochemistry. In this study three different hypotheses were tested to gain more insight into biological and ecological processes that influence the resultant composition of stable isotopes of oxygen (δ18O) in the shells of planktonic foraminifera. These hypotheses were related to the shell size; the differences in isotopic composition between the final chamber and the remaining shell; and the differences between different species. Shells of Trilobatus sacculifer, Globigerinoides ruber white and Neogloboquadrina dutertrei were picked from the top of multi-core GS07- 150-24, of modern age, offshore of north-eastern Brazil (3°46.474′ S, 37°03.849′ W) and analysed for single-shell and single-chamber stable isotope analysis. We show that the mean value of δ18O of the final chambers (δ18OF ) is 0.2%±0.4% (1ω) higher than the mean value δ18O of the test minus the final chamber (δ18O<F ) of T. sacculifer. The formation of the final chamber happens at temperatures that are approximately 1 °C cooler than the chambers formed prior, suggesting both ontogenetic depth migration to deeper water and a potential offset from the surface signal. Furthermore, we show that there is no statistical difference in the δ18Osacculifer values of shells of three different size classes of T. sacculifer, although the pattern between the different size classes indicates depth migration during the life and growth of T. sacculifer. Comparison of vital effect corrected δ18Oshell between T. sacculifer, G. ruber white and N. dutertrei suggests that G. ruber has a slightly shallower depth habitat (∼ 90-120 m) compared to the other two species (∼ 100-130 m). Disentangling depth vs. seasonal habitat is complicated given the commonality between isotope values from similar depths but different seasons; for instance, the same average isotope value will have a shallower depth habitat in May than September. Calculation of seasonal-depth habitat was therefore tested. Our results highlight the complicated nature of interpreting oxygen isotopes even for the modern record.