Achieving a superior strength-ductility combination for fcc single-phase high entropy alloys (HEAs) is challenging. The present work investigates the in-situ synthesis of Fe49.5Mn30Co10Cr10C0.5 interstitial solute-strengthened HEA containing 0.5 wt.% Nb (hereafter referred to as iHEA-Nb) using laser melting deposition (LMD), aiming at simultaneously activating multiple strengthening mechanisms. The effect of Nb addition on the microstructure evolution, mechanical properties, strengthening and deformation mechanisms of the as-deposited iHEA-Nb samples was comprehensively evaluated. Multiple levels of heterogeneity were observed in the LMD-deposited microstructure, including different grain sizes, cellular subgrain structures, various carbide precipitates, as well as elemental segregation. The incorporation of Nb atoms with a large radius leads to lattice distortion, reduces the average grain size, and increases the types and fractions of carbides, aiding in promoting solid solution strengthening, grain boundary strengthening, and precipitation strengthening. Tensile test results show that the Nb addition significantly increases the yield strength and ultimate tensile strength of the iHEA to 1140 and 1450 MPa, respectively, while maintaining the elongation over 30 %. Deformation twins were generated in the tensile deformed samples, contributing to the occurrence of twinning-induced plasticity. This outstanding combination of strength and ductility exceeds that for most additively manufactured HEAs reported to date, demonstrating that the present in situ alloying strategy could provide significant advantages for developing and tailoring microstructures and balancing the mechanical properties of HEAs while avoiding conventional complex thermomechanical treatments. In addition, single-crystal micropillar compression tests revealed that although the twining activity is reduced by the Nb addition to the iHEA, the micromechanical properties of grains with different orientations were significantly enhanced.