A PbS/PbSe nano-semiconductor co-doped silica fiber (PbS/PbSe DSF) was prepared and the enhancement mechanism of fluorescence and gain characteristics were investigated. PbS/PbSe DSF showed the broadband luminescence ranging from 1000 nm to 1380 nm, and the fluorescence intensity at 1160 nm and 1300 nm was enhanced by 15.8 dB and 14.6 dB, respectively. The on–off gain of PbS/PbSe DSF is 13.7–18.9 dB in the range of 1100–1350 nm and the net gain is 10.7–14.5 dB under 980 nm pumping. Simultaneously, based on density functional theory (DFT), local structural models of PbS/PbSe co-doped fiber materials were built, and the study showed that the introduction of PbSe increased the absorption at 969 nm and the corresponding emission in the range of 1100–1400 nm. Moreover, microstructural characterization showed the presence of high-density nanoparticle distribution in the core of PbS/PbSe DSF, indicating that the PbSe co-doping may have changed the local field environment of PbS deposition and increased the crystallinity, which finally improved the fiber performance. The results indicate that the proposed PbS/PbSe DSF is a potential gain material with important research value in areas such as active fiber amplifiers.