Development of metal nanoparticle immunoconjugates for correlative labeling in light and electron microscopy and as active targeted delivery systems

Combination of immunolabeling and targeted delivery using antibodies and nanoparticles is the focus of my thesis studies. There are two goals of employing nanoparticles and antibodies: firstly to correlatively detect multiple epitopes in biological samples in light (LM) and electron microscopy (EM) using fluorescent dyes with different excitation and emission wavelengths for LM and similarly sized colloidal nanoparticles of different elemental composition for EM; secondly, to develop a targeted delivery system consisting of colloidal gold coated iron nanoparticles that are conjugated to specific antibodies against surface antigens of target cells.;The necessity to observe biological samples at the molecular and submolecular levels has motivated development of labels to be used both in LM and EM. However, when LM and EM labels are conjugated to one antibody, the fluorescent signal is quenched because colloidal metal nanoparticles in the proximity of fluorescent dyes might absorb emission signal from fluorescent dyes. Increasing the distance between the colloidal metal nanoparticles and fluorescent dyes by applying primary and secondary IgG offers a promising approach for labels in correlative immunolabeling studies. Multiple correlative immunolabeling was studied on skeletal muscle tissue and platelets, employing two different colloidal metal nanoparticles made of gold and palladium, and two different fluorescent dyes with different excitation and emission wavelengths.;The second part of the thesis will focus on colloidal iron and gold coated iron nanoparticles in a targeted delivery system. Gold coated iron nanoparticles (core shell nanoparticles) are conjugated to antibodies specific against surface antigens of prostate cancer cells. Antibodies in this case act as delivery moieties. Once targeting was achieved, core shell nanoparticles were subjected to an oscillating magnetic field (OMF). The nanoparticles produced heating due to hysteresis loss. The heat was only produced in proximity of the nanoparticles and was sufficient to destroy cell plasma membranes of cancer cells. The results show that it is not required for nanoparticles to be internalized. The most important findings indicate that type and concentration of antibodies used are crucial factors. It is shown that in co-cultured cell lines, the target cells could be selectively destroyed using this approach.