The accumulation of ⁴He in groundwater can be a powerful tool in hydrogeologic investigations. However, the use of ⁴He often suffers from disagreement or uncertainty related to in situ and external sources of ⁴He. In situ sources are quantified by several methods, while external sources are often treated as calibration parameters in modeling. We present data from direct laboratory measurements of ⁴He release from sediments and field data of dissolved ⁴He in the Mahomet Aquifer, a well‐studied buried‐valley aquifer in central Illinois. The laboratory‐derived accumulation rates (0.13–0.91 μcm³ STP kg_water⁻¹ yr⁻¹) are 1–2 orders of magnitude greater than the accumulation rates based on the U and Th concentrations of the sediments (0.004–0.009 μcm³ STP kg_water⁻¹ yr⁻¹). The direct measurement of accumulation rates are more consistent with dissolved concentrations of ⁴He in the groundwater. We suggest that the direct measurement method is applicable in a variety of hydrogeologic settings. The patterns of accumulation of ⁴He are consistent with the conceptual model of flow in the aquifer based on hydraulic and geochemical evidence and show areas where in situ production and external sources of ⁴He are dominant. In the southwestern part of the study area, Ne concentrations are less than atmospheric solubility, indicating gases have been lost from the groundwater. Available evidence indicates that the gases are lost as groundwater passes by pockets of CH₄ in glacial deposits overlying the aquifer. However, the external flux from the underlying bedrock appears to dominate the accumulation of radiogenic ⁴He in the aquifer in the southwestern part of the study area, and the loss or gain of helium as groundwater passes through the overlying sediments is minor in comparison.