Initiation of pathways that lead to proliferation and chemoresistance by Toll-like receptors (TLRs) is an important factor in cancer progression. Here, we show the response of human cancer cells to TLR signaling inevitably linked to tumor biology. The approach is based on tailored multifunctional magnetic nanoparticles equipped with pathogen-derived ligands (CpG) functioning as TLR agonists (molecular component) to investigate the impact of transcription factor immune activation on human cancer cells. Magnetic nanoparticles (MnO and bifunctional Au-MnO) particles were covalently coated with a multifunctional polymer, displaying no cytotoxicity, to being able to enter cells while carrying foreign DNA (unmethylated CpG) to recognize intracellular TLR 9. Both, the particle and the nucleic acid are tagged with fluorescent markers for simultaneous visualization inside the cell. Apart from optical imaging, the magnetism of the particles also allows magnetic resonance imaging of organisms.