In this study, the interaction of Holmium (Ho) complex including 2, 9-dimethyl-1,10-phenanthroline, also it called Neocuproine (Neo), [Ho(Neo)Cl.HO], as fluorescence probe with Fish-Salmon-DNA (FS-DNA) is studied during experimental investigations. Multi-spectroscopic methods are utilized to determine the affinity binding constants (K) of complex-FS-DNA. It is found that fluorescence of Ho-complex is strongly quenched by the FS-DNA through a static quenching procedure. Under optimal conditions in Tris(trishydroxymethyl-aminomethane)-HCl buffer at 25 °C with pH≈7.2, intrinsic binding constant K of Ho-complex is 6.12 ± 0.04 × 10 M. Also the binding site number and Stern-Volmer quenching constant are calculated. There are different approaches, including iodide quenching assay, salt effect and thermodynamical assessment to determine the features of the binding mode between Ho-complex and FS-DNA. Also, the parent and starch and lipid nanoencapsulated Ho-complex, as potent antitumor candidates, were synthesized. The main structure of Ho-complex is maintained after encapsulation using starch and lipid nanoparticles. MTT method was used to assess the anticancer properties of Ho-complex and its encapsulated forms on human cancer cell lines of human lung carcinoma cell line and Breast cancer cell line. In conclusion, these compounds could be considered as new antitumor candidates.