Alloying copper into 316L stainless steel (316L) can enhance its antibacterial activity. In the present study, bi-metallic structures composed of 316L and 316L-10 wt.% Cu (316L-10Cu) were manufactured using PBF-LB. The microstructure and chemical homogeneity/gradients were analyzed in both the 316L-10Cu region and the 316L/316L-10Cu interface. First, cylinders of pure 316L with a diameter of 5 mm and a height of 0.7 mm were built on a 316L substrate. Subsequently, a 10 mm high section of 316L-10Cu was manufactured on top of 316L. The laser power and hatch distance were varied from 180 to 195 W and from 0.05 to 0.11 mm, respectively, while the powder layer thickness (50 μm) and laser scanning speed (600 mm/s) were kept constant. Increasing the volumetric energy density changed the melting mode from conductive to the keyhole mode, which improved mixing and promoted a more homogeneous distribution of Cu in 316L. However, irregular and large lack-of-fusion defects were observed at lower hatch spacing, attributed to increased spatter formation under high volumetric energy density. Furthermore, 316L-10Cu exhibited hot cracking along Cu-rich solidification grain boundaries. The interfacial region showed a gradual variation in Cu concentration, while regions containing less than 7.5 wt.% Cu were free from cracking. Copper alloying resulted in about a 16% reduction in the hardness. Hence, 316L can be functionalized for antibacterial activity by alloying with approximately 7.5 wt.% Cu, despite the associated reduction in hardness.