Synthesis and Characterization of Functional Nanofilm-Coated Live Immune Cells

Layer-by-layer (LbL) assembly techniques have been extensively studied in cell biology because of their simplicity of preparation and versatility. The applications of the LbL platform technology using polysaccharides, silicon, and graphene have been investigated. However, the applications of the above-mentioned technology using living cells remain to be fully understood. This study demonstrates a living cell-based LbL platform using various types of living cells. In addition, it confirms that the surplus charge on the outer surface of the coated cells can be used to bind the target protein. We develop a living cell-based LbL platform technology by stacking layers of hyaluronic acid (HA) and poly-l-lysine (PLL). The HA/PLL stacking results in three bilayers with a thickness of 4 ± 1 nm on the cell surface. Furthermore, the multilayer nanofilms on the cells are completely degraded after 3 days of the application of the LbL method. We also evaluate and visualize three bilayers of the nanofilm on adherent (AML-12 cells)-, nonadherent (trypsin-treated AML-12 cells)-, and circulation type [peripheral blood mononuclear cells (PBMCs)] cells by analyzing the zeta potential, cell viability, and imaging via scanning electron microscopy and confocal microscopy. Finally, we study the cytotoxicity of the nanofilm and characteristic functions of the immune cells after the nanofilm coating. The multilayer nanofilms are not acutely cytotoxic and did not inhibit the immune response of the PBMCs against stimulant. We conclude that a two bilayer nanofilm would be ideal for further study in any cell type. The living cell-based LbL platform is expected to be useful for a variety of applications in cell biology

Enhanced Doubly Activated Dual Emission Fluorescent Probes for Selective Imaging of Glutathione or Cysteine in Living Systems

The development of novel fluorescent probes for monitoring the concentration of various biomolecules in living systems has great potential for eventual early diagnosis and disease intervention. Selective detection of competitive species in biological systems is a great challenge for the design and development of fluorescent probes. To improve on the design of fluorescent coumarin-based biothiol sensing technologies, we have developed herein an enhanced dual emission doubly activated system (<b>DACP-1</b> and the closely related <b>DACP-2</b>) for the selective detection of glutathione (GSH) through the use of one optical channel and the detection of cysteine (Cys) by another channel. A phenylselenium group present at the 4-position completely quenches the fluorescence of the probe via photoinduced electron transfer to give a nonfluorescent species. Probes are selective for glutathione (GSH) in the red region and for cysteine/homocysteine (Cys/Hcy) in the green region. When they were treated with GSH, <b>DACP-1</b> and <b>DACP-2</b> showed strong fluorescence enhancement in comparison to that for closely related species such as amino acids, including Cys/Hcy. Fluorescence quantum yields (Φ_F) increased for the red channel (<0.001 to 0.52 (<b>DACP-1</b>) and 0.48 (<b>DACP-2</b>)) and green channel (Cys) (<0.001 to 0.030 (<b>DACP-1</b>) and 0.026 (<b>DACP-2</b>)), respectively. Competing fluorescent enhancements upon addition of closely related species were negligible. Fast responses, improved water solubility, and good cell membrane permeability were all properly established with the use of <b>DACP-1</b> and <b>DACP-2</b>. Live human lung cancer cells and fibroblasts imaged by confocal microscopy, as well as live mice tumor model imaging, confirmed selective detection

Facile Solvothermal Preparation of Monodisperse Gold Nanoparticles and Their Engineered Assembly of Ferritin–Gold Nanoclusters

Herein, we report a quick and simple synthesis of water-soluble gold nanoparticles using a HAuCl₄ and oleylamine mixture. Oleylamine serves as a reduction agent as well as a stabilizer for nanoparticle surfaces. The particle sizes can be adjusted by modulating reaction temperature and time. Solvothermal reduction of HAuCl₄ with oleylamine can be confirmed by measuring the product in Fourier transform infrared (FTIR) spectroscopy. The plasmon band shifting from yellow to red confirms a nanosized particle formation. Amide bonds on the surface of the nanoparticles formed hydrogen bonds with one another, resulting in a hydrophobic monolayer. Particles dispersed well in nonpolar organic solvents, such as in hexane or toluene, by brief sonication. Next, we demonstrated the transfer of gold nanoparticles into water by lipid capsulation using 1-myristoyl-2-hydroxy-<i>sn</i>-glycero-3-phosphocholine (MHPC), 1,2-distearoyl-<i>sn</i>-glycero-3-phosphoethanolamine-<i>N</i>-(methoxy polyethylene glycol)-2000 (DPPE-PEG2k), and 1,2-dioleoyl-<i>sn</i>-glycero-3-<i>N</i>-{5-amino-1-carboxypentyl}­iminodiacetic acid succinyl nickel salt [DGS-NTA­(Ni)]. The particle concentration can be obtained using an absorbance in ultraviolet–visible (UV–vis) spectra (at 420 nm). Instrumental analyses using transmission electron microscopy (TEM), energy-dispersive X-ray (EDX) analysis, dynamic light scattering (DLS), and FTIR confirmed successful production of gold nanoparticles and fair solubility in water. Prepared gold particles were selectively clustered via engineered ferritin nanocages that provide multiple conjugation moieties. A total of 5–6 gold nanoparticles were clustered on a single ferritin nanocage confirmed in TEM. Reported solvothermal synthesis and preparation of gold nanoclusters may serve as an efficient, alternate way of preparing water-soluble gold nanoparticles, which can be used in a wide variety of biomedical applications

Mesoporous Silica-Coated Hollow Manganese Oxide Nanoparticles as Positive <i>T</i>₁ Contrast Agents for Labeling and MRI Tracking of Adipose-Derived Mesenchymal Stem Cells

Mesoporous silica-coated hollow manganese oxide (HMnO@mSiO₂) nanoparticles were developed as a novel <i>T</i>₁ magnetic resonance imaging (MRI) contrast agent. We hypothesized that the mesoporous structure of the nanoparticle shell enables optimal access of water molecules to the magnetic core, and consequently, an effective longitudinal (<i>R</i>₁) relaxation enhancement of water protons, which value was measured to be 0.99 (mM⁻¹s⁻¹) at 11.7 T. Adipose-derived mesenchymal stem cells (MSCs) were efficiently labeled using electroporation, with much shorter <i>T</i>₁ values as compared to direct incubation without electroporation, which was also evidenced by signal enhancement on <i>T</i>₁-weighted MR images in vitro. Intracranial grafting of HMnO@mSiO₂-labeled MSCs enabled serial MR monitoring of cell transplants over 14 days. These novel nanoparticles may extend the arsenal of currently available nanoparticle MR contrast agents by providing positive contrast on <i>T</i>₁-weighted images at high magnetic field strengths