Composite nanoparticles with magnetic core and fluorescent shell were facilely prepared by the layer-by-layer deposition of conjugated polyelectrolytes over the negatively charged nanoparticles (NPs) of superparamagnetic iron oxide. The alternate assembly of cationic and anionic fluorescent polyelectrolytes leads to reversal in the sign of ζ-potentials. The even numbers of adsorption layer corresponding to the anionic polyelectrolyte (PFS) have negative values (−13 to −24 mV), whereas odd numbers of coating relative to the cationic polyelectrolyte (PFN) have positive values (26 to 28 mV). The composite nanoparticles can respond to both external magnetic field and ultraviolet light excitation. Förster resonance energy transfer (FRET) between oppositely charged polyelectrolytes (PFN and ThPFS) layers was also found, indicating dense packing of the polymer coatings. The fluorescence of the positively charged nanoparticles (NPs/PFN) can be quenched with very high efficiency by a small molecule anionic quencher Fe(CN)6 4−, while the same quencher has far less effect on the fluorescence of the negatively charged nanoparticles (NPs/PFN/PFS).