Shells of the northern and southern quahogs ( Mercenaria mercenaria and Mercenaria campechiensis, respectively) from paleontological and archaeological deposits have been used to reconstruct climate, environmental conditions, and ecological relationships. Seasonal fluctuations in temperature and salinity are recorded in shell carbonate as variations in δ 18O and δ 13C values and Sr : Ca ratios; however, these environmental proxies have not yet been calibrated for M. campechiensis. Previous geochemical studies of quahog shells focused on the outer prismatic layer as an ecological archive. The outer layer is often not preserved in fossil and archaeological shells. Moreover, recent innovations in microsampling techniques allow high-resolution sampling of the middle cross-lamellar layer. This study tests whether outer prismatic and middle cross-lamellar layers record similar variations in δ 18O and δ 13C values and Sr : Ca ratios in modern shells of M. campechiensis and whether profiles are similar among individuals. The last 2 years of shell growth from three individuals (6, 7, and 9 years old) collected alive near Bokeelia, southwest Florida, were microsampled for geochemical analyses. δ 18O varies seasonally relative to translucent (summer) and opaque (winter) growth increments. Variation of δ 18O from outer and middle layers are identical within and among shells; however, the oldest individual did not preserve the most positive values represented in the two younger shells. δ 13C from both layers are similar within shells, although the outer layer is more variable and the range of values varies somewhat among individuals. Sr : Ca ratios from the outer and middle layers within shells do not track each other well, and profiles among individuals are different. To evaluate whether a seasonal signal was present, Sr : Ca ratios were compared to δ 18O values. There was no co-variation within the outer layer in any of the shells; therefore, this layer should be avoided when using Sr : Ca ratios as a temperature proxy. The middle layer from the 6-year-old shell showed no co-variation, the 7-year-old shell exhibited a moderate negative co-variation ( r 2 = 0.55, p < 0.01), and the 9-year-old shell reflected a strong negative co-variation ( r 2 = 0.72, p < 0.01). These findings suggest that Sr : Ca ratios from the middle layer may serve as a proxy for temperature only in older individuals with slower growth rates. Either shell layer can be sampled for assessing seasonality using δ 18O, and environmental interpretations made using δ 13C values should be treated with caution.