In this work, citric acid-based quantum dots (CA-QDs) as a novel and safe crosslinked agent was applied in different feeding ratios (5–15 wt%) to synthesize carboxymethyl cellulose/polyvinyl alcohol (CMC/PVA) nanofibers (NFs) for the first time. Colistin (CL) as an antibacterial agent was also loaded (2 w/w%) during the synthesizing process of CMC/PVA electrospun NFs to trigger antimicrobial properties. The morphological, hydrophilic, and mechanical properties of the prepared NFs were fully investigated with different techniques. The electrospun NFs with crosslinking ratios of 10 wt% CA-QDs revealed appropriate mechanical properties. According to cell culture data, the prepared NFs demonstrated good cytocompatibility against HFF-1 cells (over 80% cell viability). Remarkably, CL-loaded NFs showed desired antibacterial efficacy against S. aureus, E. coli, K. pneumoniae, and P. aeruginosa with 1.0–1.4, 1.3–1.4, 0.8–1.0, and 1.3–1.5 cm inhibition zones, respectively. These outcomes suggested that the fabricated NFs can be useful as wound healing scaffolds.