In this study, a system to measure leaf internal CO2 (Ci) was incorporated into an open gas exchange system (LI-COR, Lincoln, NE, USA). The Ci was directly measured with a cup attached to the abaxial surface of sunflower (Helianthus annuus L.) leaves with open stomata while normal CO2 and water vapor exchange through the same section of adaxial surface was simultaneously detected. The potential problems in the system, namely bulk air flow through the leaf, diffusion leaks, and change in the CO2 gradient inside the leaf, were examined with the aim to apply the system to measure net photosynthesis at various Ci (i.e. A Ci curves). A micro blower constantly circulated the air in a loop without pressure pulses or bulk air movement through the leaf. The measured Ci (Ci(m)) generally followed the external CO2 as much as the calculated one (Ci(c)). There was close agreement between the Ci(m) and the Ci(c) particularly at low Ci, and the diffusion leak hardly affected the relationship between the two. Despite possible alterations of leaf properties by cup attachment, the direct measurement is expected to cast a new light on leaf gas exchange.