A unique and potent protein binding nature of liposome containing polyethylenimine and polyethylene glycol: a nondisplaceable property.

Most of the currently available targeting vectors are produced via the linkage of targeting molecules. However, the coupling process is complicated, and the covalent linkage may attenuate the activity of certain targeting molecules. In this study, we have developed a cationic liposome complexed with polyethylenimine and polyethylene glycol polymers (LPPC) that can capture various proteins without covalent conjugation. Characterizations of prepared LPPC revealed that the maximal-binding capacity was about 170 µg of bovine serum albumin to 40 µg of sphere-shaped LPPC (180 nm). The proteins were essentially located at or near the surface when analyzed by atomic force or transmission electron microscopy. We demonstrate that polyethylenimine was an essential component to bind the proteins. Upon the saturation of captured proteins, a given protein could not be displaced by other additional proteins and still retained its biological activity. Using a variety of functional proteins, we show some typical examples of the utility of incorporated beta-glucuronidase and antibodies onto the LPPC. The beta-glucuronidase can be used for the study of antigen-antibody interactions, whereas in studies with the antibody complex, we used anti-CD3 as an agonist to stimulate the proliferation of peripheral blood mononuclear cells via a receptor-mediated mechanism and anti-VEGFR for cell staining. In conclusion, the prepared LPPC can provide a platform to capture biologically and biochemically functional proteins on its surface for various applications, such as cell signaling, cell profiling, noncovalent enzyme-linked immunoassays, and others not mentioned.

Multifunctional core-shell polymeric nanoparticles for transdermal DNA delivery and epidermal Langerhans cells tracking.

Skin is a highly immune-reactive tissue containing abundant antigen-presenting cells such as Langerhans cells (LCs), and thus is a favorable site for DNA immunization. This study developed a multifunctional core-shell nanoparticle system, which can be delivered transdermally into the epidermis via a gene gun, for use as a DNA carrier. The developed nanoparticles comprised a hydrophobic PLGA core and a positively-charged glycol chitosan (GC) shell. The core of the nanoparticles was used to load fluorescent quantum dots (QDs) for ultrasensitive detection of Langerhans cell migration following transdermal delivery, while a reporter gene was electrostatically adsorbed onto the GC shell layer of the nanoparticles. Results of fluorescence spectrophotometry, transmission electron microscopy, energy dispersive X-ray analysis, and X-ray diffraction measurement confirmed that the prepared nanoparticles had a core-shell structure with QDs in their core area. The surface charge of nanoparticles depended strongly on pH environment, enabling the intracellular release of the loaded DNA via a pH-mediated mechanism. Using a mouse model, this study demonstrated that bombardment of nanoparticles transfected DNA directly into LCs present in the epidermis; the transfected LCs then migrated and expressed the encoded gene products in the skin draining lymph nodes. These observation results suggest that the developed nanoparticle system is suitable for monitoring and fine-tuning important functional aspects of the immune system, in conjunction with the loaded fluorescence, and thus has potential for use in immunotherapy and vaccine development.

The characteristics, biodistribution, magnetic resonance imaging and biodegradability of superparamagnetic core-shell nanoparticles.

An efficient contrast agent for magnetic resonance imaging (MRI) is essential to enhance the detection and characterization of lesions within the body. In this study, we described the development of biodegradable nanoparticles with a core-shell structure to formulate superparamagnetic iron oxide (CSNP-SPIO) for MRI. The developed nanoparticles were composed of a hydrophobic PLGA core and a positively-charged glycol chitosan shell. The results obtained by transmission electron microscopy, energy dispersive X-ray analysis, electron energy loss spectroscopy, and X-ray diffraction measurement confirmed that the prepared nanoparticles had a core-shell structure with SPIO in their core area. No aggregation of nanoparticles was observed during storage in water, as a result of the electrostatic repulsion between the positively-charged nanoparticles. The magnetic properties of nanoparticles were examined by a vibrating sample magnetometer and a superconducting quantum interference device; the results showed that the superparamagnetism of SPIO was preserved after the CSNP-SPIO formulation. In tracking their cellular internalization pathway, we found that CSNP-SPIO accumulated in lysosomes. In the biodistribution study, a high level of radioactivity was observed in the liver shortly after administration of the (99m)Tc-labeled CSNP-SPIO intravenously. Once taken up by the liver cells, the liver turned dark on T(2)* images. Following cellular internalization, CSNP-SPIO were broken down gradually; therefore, with time increasing, a significant decrease in the darkness of the liver on T(2)* images was found. The aforementioned results indicate that the developed CSNP-SPIO can serve as an efficient MRI contrast agent and could be degraded after serving their imaging function.

Phase TEM for biological imaging utilizing a Boersch electrostatic phase plate: theory and practice.

A Boersch electrostatic phase plate (BEPP) used in a transmission electron microscope (TEM) system can provide tuneable phase shifts and overcome the low contrast problem for biological imaging. Theoretically, a pure phase image with a high phase contrast can be obtained using a BEPP. However, a currently available TEM system utilizing a BEPP cannot achieve sufficiently high phase efficiency for biological imaging, owing to the practical conditions. The low phase efficiency is a result of the blocking of partial unscattered electrons by BEPP, and the contribution of absorption contrast. The fraction of blocked unscattered beam is related to BEPP dimensions and to divergence of the illumination system of the TEM. These practical issues are discussed in this paper. Phase images of biological samples (negatively stained ferritin) obtained by utilizing a BEPP are reported, and the phase contrast was found to be enhanced by a factor of approximately 1.5, based on the calculation using the Rose contrast criterion. The low gain in phase contrast is consistent with the expectation from the current TEM/BEPP system. A new generation of phase TEM utilizing BEPP and designed for biological imaging with a high phase efficiency is proposed.

Bandgap mapping for III-V quantum well by electron spectroscopy imaging.

The concept of 'bandgap mapping' was proposed originally to map the inhomogeneity of band energy in III-V semiconductors with a spatial resolution of a few nanometres in a scanning transmission electron microscope with the focus beam mode. In this paper, several techniques were developed to demonstrate the possibility to map the distribution of bandgap energies for GaN/AlN quantum-well structures using electron spectroscopy imaging (ESI). The phase correlation function was used to register different energy-loss images among ESI series with an accuracy of 1 pixel. The energy dispersion of ESI series was improved by a fast Fourier transform interpolation method. An iterative multivariable least square algorithm was derived to refine the fitting of the single scattering distribution to an analytic form of the density of states function a(E - E(g))(0.5). The inhomogeneity of the band energy of the quantum well can be revealed from the band-energy map. A threshold filter method is applied to estimate the average value and SD of the bandgap energy from barrier and well regions in the energy map. The average bandgap energy of AlN and GaN is determined to be 5.62 +/- 0.35 and 3.87 +/- 0.36 eV, respectively. The effect of delocalization on the accuracy of band-energy determination is discussed. The 2sigma(E(g)) accuracy of this analysis is comparable to half of the energy resolution of the ESI experiment.