S. A. Staroverov

Use of a synthetic foot-and-mouth disease virus peptide conjugated to gold nanoparticles for enhancing immunological response

Foot-and-mouth disease is an acute, highly contagious infection of domestic and wild cloven-hoofed animals, which can be transmitted to humans. In many cases, the existing vaccines are not quite effective. The purpose of this study was to test the possibility of using gold nanoparticles as an antigen carrier and an adjuvant. The immunogenic properties of gold nanoparticles were assessed by conjugating the particles to a synthetic peptide of the VP1 capsid protein of the foot-and-mouth disease virus. The resulting conjugate (with or without the use of complete Freund’s adjuvant), a commercial vaccine, and the native peptide served to immunize guinea pigs. The titer and sensitivity of the raised antibodies were maximal for the combination comprising the nanoparticle–peptide conjugate and complete Freund’s adjuvant. Antibody biosynthesis was accompanied by increased production of proinflammatory cytokines (especially IFN-γ) and by stimulation of the respiratory activity of peritoneal macrophages. The use of gold nanoparticles as a hapten carrier enhanced the immune response even when complete Freund’s adjuvant was not used.

Analytical and Theranostic Applications of Gold Nanoparticles and Multifunctional Nanocomposites

Gold nanoparticles (GNPs) and GNP-based multifunctional nanocomposites are the subject of intensive studies and biomedical applications. This minireview summarizes our recent efforts in analytical and theranostic applications of engineered GNPs and nanocomposites by using plasmonic properties of GNPs and various optical techniques. Specifically, we consider analytical biosensing; visualization and bioimaging of bacterial, mammalian, and plant cells; photodynamic treatment of pathogenic bacteria; and photothermal therapy of xenografted tumors. In addition to recently published reports, we discuss new data on dot immunoassay diagnostics of mycobacteria, multiplexed immunoelectron microscopy analysis of Azospirillum brasilense, materno-embryonic transfer of GNPs in pregnant rats, and combined photodynamic and photothermal treatment of rat xenografted tumors with gold nanorods covered by a mesoporous silica shell doped with hematoporphyrin.

Immunogenic Properties of Colloidal Gold

We studied the capacity of colloidal gold for enhancing specific and nonspecific immune response in laboratory animals (rabbits, rats, and mice) immunized with antigens of various nature. The antibody titers obtained with colloidal gold as a carrier were higher as compared to the standard immunization techniques (free antigen or its combination with Freunds adjuvant). Application of colloidal gold also enhanced nonspecific immune responses, such as lysozyme concentration in the blood, activity of the complement system proteins, as well as phagocytic and bactericidal activities. The antibodies were tested by immunodot assay using gold markers. Immunization of the animals with colloidal gold conjugates with haptens or complete antigens (without other adjuvants) was shown to induce the production of highly active antibodies. In addition, the amount of antigen used for animal immunization with colloidal gold was an order of magnitude lower, compared to immunization with complete Freunds adjuvant. This fact can be evidence for adjuvant properties of colloidal gold proper.

Quantitative cell bioimaging using gold-nanoshell conjugates and phage antibodies.

The authors describe a quantitative evaluation of the efficacy of cell labeling with plasmon-resonant light-scattering nanoparticles used as contrast agents for dark-field microscopy imaging. The experimental model is based on the biospecific labeling of pig embryo kidney (SPEV) cells with primary phage antibodies, followed by the dark-field microscopic visualization using conjugates of silica/gold nanoshells with secondary rabbit antiphage antibodies. To quantify nanoparticle binding, the authors introduce the labeling-efficacy factor (LEF) which is equal to the ratio of the bound-particle pixels per cell to the total number of pixels occupied by the cell. The LEF is calculated by an imaging-analysis algorithm based on the freely available ImageJ Java-based processing code. In terms of the LEF, a distinct difference was found between intact, nonspecifically labeled, and biospecifically labeled cells.

Effect of gold nanoparticles on the respiratory activity of peritoneal macrophages

This work was undertaken to investigate the effect of antigen-conjugated gold nanoparticles on the functional activity of cells of the reticuloendothelial system. Experiments using rat and mouse peritoneal cells assessed the dynamics of the total respiratory activity of macrophages and the activity of mitochondrial dehydrogenases on exposure to colloidal gold conjugates of high- and lowmolecular-weight antigens. Both gold nanoparticles and their antigen conjugates stimulated the respiratory activity of the macrophages and the activity of macrophage mitochondrial enzymes. This stimulation may be an essential factor determining the adjuvant properties of colloidal gold, found by us in prior work.

Adjuvant Properties of Gold Nanoparticles

This review summarizes known data and the results of our own investigations into the application of gold nanoparticles as an antigen (AG) carrier and as an adjuvant in immunization in the in vivo preparation of antibodies (ABs). We have developed a technology for the production of ABs against various AGs by using colloidal gold as a carrier and as an adjuvant. The repeated injection of colloidal gold (CG)-AG conjugates (with or without the use of Freund’s complete adjuvant) into animals yielded specific high-titer ABs for a variety of AGs with no concomitant ABs. Gold nanoparticles used as an AG carrier activated the phagocytic activity of macrophages and influenced the functioning of lymphocytes, which apparently may be responsible for their immunomodulating effect. Thus, gold nanoparticles can facilitate the synthesis of ABs in rabbits, rats, and mice, in particular, by reducing the amount of required AG when compared to immunization using Freund’s complete adjuvant.

New Types of Nanomaterials: Powders of Gold Nanospheres, Nanorods, Nanostars, and Gold-Silver Nanocages

We describe a new strategy for the fabrication of plasmonic nanopowders (dried gold or composite nanoparticles) by using wet chemical nanoparticle synthesis, PEG-SH functionalization, and freeze drying. This strategy is illustrated by the successful production of four plasmonic nanopowders, including gold nanorods, nanostars, gold-silver nanocages, and gold nanospheres. The dried nanoparticles can be stored for a long time under ordinary conditions and then easily dissolved in water at a desired concentration without heating, intense sonication, or other harsh treatments. According to the MTT assay with SPEV-2 cells, gold nanorod powder is not toxic at concentrations up to 100 mg/L. Redispersed samples maintain the plasmonresonant properties of the parent colloids and do not form aggregates. These properties make pegylated freeze-dried gold nanoparticles (GNPs) applicable in various fields of biomedicine.

Immunostimulatory Effect of Gold Nanoparticles Conjugated with Transmissible Gastroenteritis Virus

Animals were immunized with transmissible gastroenteritis virus conjugated with gold nanoparticles. The resultant antibodies had a higher titer than antibodies produced in response to native virus. Immunization with the antigen-colloidal gold complex led to a significant increase of the peritoneal macrophages respiratory activity and of plasma IFN-γ level in immunized animals.

Plasmon-Resonant Gold Nanoparticles with Variable Morphology as Optical Labels and Drug Carriers for Cytological Research

In this work, two types of nanocomposites, silica-coated nano-sea-urchins and silica-coated gold nanostars, were fabricated. CTAB-coated nano-sea-urchins with an average size of about 100 nm demonstrate an absorption peak near 600-700 nm and stability in aqueous suspension. CTAB was exchanged with m-PEG-SH by an intermediate PEG layer. A layer of silica was synthesized on the nano-sea-urchins surface with thickness of about 20 nm. Nanostars with an average size of about 60 nm with a number of thin sharp branches were fabricated and functionalized by PVP to improve their stability. PVP-coated nanostars were used in optical coherence tomography experiments to show their contrasting properties. After silica-coating, stable and monodispersed nanoparticles with silica shell thickness about 60 nm were obtained. Nontoxicity of the silica-coated nanostars at least until the concentration of nanoparticles about 400 μg/mL was showed by fluorescent cell viability assay using propidium iodide. Extinction coefficient of the gold nanostars and nanocomposites was estimated by a spectrophotometer system in collimated transmission regime.

Obtainment of Polyclonal Antibodies to Clenbuterol with the Use of Colloidal Gold

The effectiveness of an in vivo synthesis of antibodies to clenbuterol with the use of gold nanoparticles as a carrier was evaluated. For comparison, conjugates of clenbuterol with bovine serum albumin were used in immunization. The serum titer was determined by an ELISA. The antibodies were tested by an immunodot assay with immunogold markers. With both techniques we obtained specific and relatively high-titer antibodies to clenbuterol. It was found that the antibodies obtained with the use of gold nanoparticles were not inferior in titer to those obtained to the conjugates of clenbuterol with the protein but surpassed them in specificity.