This paper presents the optimal selection of cable reinforcement for concrete 3D-printed lattice beam. It also addresses key challenges associated with interlayer bonding and flexural strength of 3D printed elements. Three different types of reinforcement including 1.6 mm and 2 mm diameter rebar, (R1.6 & R2) and 1.5 mm wire (W1.5) with seven strings were evaluated with different stand-off distances of 10–20 mm. The interlayer bonding and flexural strength were investigated to determine the most effective reinforcement configuration. Results show that W1.5 reinforcement exhibited higher interlayer bonding and flexural strength as compared with R1.6 and R2. Subsequently, the optimal reinforcement configuration (W1.5) was embedded in a concrete 3D-printed lattice beam. The lattice beam was structurally evaluated using four-point testing, demonstrating an ultimate load-carrying capacity of 49.01 kN with a mid-span displacement of 2.99 mm. Moreover, it exhibited flexural failure with no premature failures such as inter-layer delamination.