Quantum dot (QD) light-emitting devices operating in non-carrier-injection (NCI) mode have attracted intense interest. Revealing the source of carriers that support the periodic electroluminescence is important because there is no injection of carriers from the external electrode. Electrons/holes generated by well-to-well multiple ionization in adjacent QDs are generally recognized as the carrier source for electroluminescence, and the stacked QD layers are necessary. In this work, NCI electroluminescence (NCI-EL) from monolayer QDs is successfully demonstrated, which cannot be properly explained by the previously proposed mechanism of multiple ionization. A working mechanism related to periodic in-well ionization is proposed, in which electrons tunnel directly from the valence band of QDs to the conduction band to form free electrons and holes. The effects of driving voltage amplitude, frequency, and QD size on the NCI-EL performance are investigated. Finite element simulation is used to clarify the ionization process. We believe this work can extend the working mechanism model of NCI-EL from QDs and provide guidance for promoting QD-based light-emitting device performance.