Poly(D,L-lactide-co-glycolide) (PLGA) nanoparticles loaded with lamivudine and coated with bovine serum albumin (BSA) were prepared via a double emulsion method. The influences of experiments parameters such as volume of inner aqueous phase, concentration of organic phase and ultrasonication time on the particle size and drug entrapment efficiency were investigated, obtaining PLGA particles with a diameter of ∼260 nm and drug entrapment efficiency of ∼35%. The particles were observed by scanning electron microscopy and transmittance electron microscopy, showing a core-shell structure. BCA assay found that 58 mg BSA was present on/in 1 g LPB particles. The loaded lamivudine showed a burst release at beginning and sustained release until 24 h in physiological conditions. Low pH could accelerate the release of lamivudine from PLGA particles, making the PLGA particles potential intelligent intracellular drug carriers. The PLGA particles were readily internalized into the human liver cells within a short time and increased gradually with the prolongation of incubation time regardless of the loading of lamivudine. The particles either resided within lysosomes or transferred to cytoplasm, but could not enter into the cell nucleus. The cell viability was not significantly influenced in the presence of the particles regardless of lamivudine encapsulation, suggesting that this kind of particles may be a good candidate for the intracellular anti-hepatitis B drug delivery.