An important tool to identify the origin of a groundwater resource is the use of isotopic signatures. Isotopic signatures give us the age of water and provide information as to the water’s origin, potential transit at geologic structures, source of salinization, and possible recharge points. The purpose of this study was to collect and analyze well samples to evaluate isotopic tracers (δ18O and tritium) in the transboundary Conejos-Médanos/Mesilla aquifer located between the US and Mexico. This new analyzed information was compared with the isotopic information available in the US Mesilla and US-MX Hueco basins generated by previous works, which described the common origin of the Hueco Bolson and Mesilla Basins aquifers. This study used isotopic analysis to validate the theory of the original formation and interconnectivity of both transboundary basins. This research presents new data of δ18O and tritium, and a comparison with previous published data from other workers, versus the known global meteoric water line (GMWL) and the Rio Grande evaporation line (RGEL). Results show that the groundwater at the transboundary aquifer features an evaporated isotopic signal, which is consistent with referenced published data that discusses the geologic history of aquifer formations at the studied area. This study is important because isotopic studies from the area were nonexistent and because isotopic data can explain recharge scenarios that relate to groundwater quality.