Figure 5.

Comparison of antibody responses, nanoparticle location, and phagocyte uptake of targeting ligand conjugated IONPs and PEG-IONPs following s.c. injections. NIR-830 dye labeled mATF or hATF peptides were conjugated to IONPs or PEG-IONPs and then injected s.c. into the front flank area of normal Balb/c mice. A. Levels of anti-ATF antibodies in the serum of the mice received mATF or hATF conjugated IONPs, without or with PEG-modification. 1 and 2 weeks following IONP injections, mouse serum samples were collected and ELISA assay was used for the detection of antibody responses. n=3 mice/group. B. Optical imaging detected location and migration of the IONPs without or with PEG modification. Optical imaging was performed 1, 2 and 7 days following the first injection using the Kodak FX In Vivo imaging system. Optical imaging signals at the injection site (pink arrows). Diffused signals in the draining lymph node areas of the mice injected with NIR-830-mATF-PEG-IONPs (blue arrows). Numbers shown are the mean signal intensities of the injection site. C. Prussian blue staining showed the presence of cells with IONPs in the injection sites, the draining lymph nodes and the spleen obtained from the mice 7 days following the last nanoparticle injection (red arrows). D. Immunohistochemical staining. Tissue sections of the injection site were stained with antibodies to CD68 (macrophages and dendritic cells) and CD83 (mature dendritic cells). CD68 and CD83 positive cells were found in the injection site (red arrows). A higher level of CD83 positive cells was detected in the tissue sections of the mice injected with PEG modified-ATF-IONPs compared with ATF-IONP injection site. Green arrows indicated the areas of CD83 positive Langerhans cells, which are skin dendritic cells.