In recent years, the application of upconversion nanoparticles, especially in vivo, has so far largely been hampered due to the strong tissue absorption of short-wavelength light below 600 nm. Therefore, it is an urgent need to carry out some new selective approaches to increase red emission efficiency. This work reports an effective strategy to enhance red upconversion emission of NaYF4:Yb3+, Er3+ nanocrystals by doping with the different concentrations of transition metal ions (Mn2+ or Fe3+). The upconversion emission spectra of the samples demonstrated that the red emissions were significantly enhanced and that different transition metal ions had a distinguishable effect on the red upconversion luminescence. Compared with the pure NaYF4:Yb3+, Er3+ nanoparticles, the red upconversion emission intensity of the nanoparticles co-doped with 10 mol% Mn2+ was increased by nearly as high as 14 times. In addition, the red emission intensity was increased by about 24 times, which was observed in the sample co-doped with 30 mol% Fe3+. The mechanism of the enhancement was discussed detailedly. Both the improvement of the nanoparticles׳ crystallinity and the redistribution of electron cloud around the Er3+ ions after introducing Mn2+ or Fe3+ were confirmed to be the origin of the improvement. The enhancement in red upconversion luminescence intensity makes the upconversion nanoparticles versatile imaging tools for diagnosis.