Folate conjugated N-trimethyl chitosan (folate-TMC) was synthesized and characterized using Fourier transform infrared (FTIR) and (1)H spectroscopy. The fluorescein isothiocyanate conjugated bovine serum albumin (FITC-BSA) loaded TMC-nanoparticle (FB-TMC-NP) and FITC-BSA loaded folate-TMC-nanoparticle (FB-f-TMC-NP) were prepared by ionic cross-linking of TMC or folate-TMC with sodium alginate. Single factor analysis method was used to optimize the formulation of nanoparticles. The encapsulating efficiencies of FB-TMC-NP and FB-f-TMC-NP produced by optimal formulation were 98.3 +/- 1.9% and 98.7 +/- 2.7% (n=3), respectively. In addition, the mean diameters of FB-TMC-NP and FB-f-TMC-NP were 184.3 +/- 8.3 nm and 176.1 +/- 5.0 nm (n = 3), respectively. Transmission electron microscope (TEM) showed that the nanoparticles were of spherical shapes. The intracellular uptake of FB-f-TMC-NP by SKOV3 cells (folate receptor overexpressing cells) was 3.7-fold more than that of FB-TMC-NP and could be inhibited by the presence of 1 mM folate in the culture medium, although there was no significant difference between the intracellular uptake of FB-f-TMC-NP in A549 cells (folate receptor-deficient cells) and that of FB-TMC-NP in the same cells. In conclusion, the enhancement of cellular uptake of FB-f-TMC-NP by SKOV3 cells in a specific way was attributed to the folate receptor-mediated endocytosis. FB-TMC-NP was a promising carrier for protein.