Two-photon excited fluorescence materials usually suffer from inefficient two-photon absorption (TPA) and nonradiative excited states. Here, upconversion fluorescence in an electron donor-acceptor (DA) exciplex doped with fluorescent emitters are systematically investigated. It has been found that the undoped DA exciplex exhibits enhancements of ～129% and ～365% in upconversion fluorescence compared to donor- and acceptor-only systems, respectively. Interestingly, photoluminescence quantum yields (PLQYs) up to ～98.65% were measured and immensely enhanced upconversion fluorescence was observed after doping various fluorescent emitters into the DA exciplex. Our results reveal the existence of two-photon excited energy harvesting in a thermally activated delayed fluorescence (TADF) DA exciplex doped with fluorescent emitters, via reverse intersystem crossing followed by rapid F?rster resonance energy transfer. Moreover, the additional gain mechanism related to intermolecular CT interaction that occurs at the TPA stage is found in the TADF DA exciplex system.