Chimeric antigen receptor (CAR)-modified T cell therapy has the potential to improve the overall survival of patients with malignancies by enhancing the effectiveness of CAR T cells. Precisely predicting the effectiveness of various CAR T cells represents one of today's key unsolved problems in immunotherapy. Here, we predict the effectiveness of CAR-modified cells by evaluating the quality of the CAR-mediated immunological synapse (IS) by quantitation of F-actin, clustering of tumor antigen, polarization of lytic granules (LGs), and distribution of key signaling molecules within the IS. Long-term killing capability, but not secretion of conventional cytokines or standard 4-hr cytotoxicity, correlates positively with the quality of the IS in two different CAR T cells that share identical antigen specificity. Xenograft model data confirm that the quality of the IS in vitro correlates positively with performance of CAR-modified immune cells in vivo. Therefore, we propose that the quality of the IS predicts the effectiveness of CAR-modified immune cells, which provides a novel strategy to guide CAR therapy.

New spherical nanostructures of titania (TiO2) have been synthesized through formation of liposome−TiO2 nanocomposites by using egg lecithin lipid as a template, and their optical properties have been investigated with regard to the dynamics of surface charge carriers and photocatalytic activities by using UV−vis and photoluminescence (PL) spectroscopic techniques. On the basis of the measurements of X-ray diffraction, transmission electron microscopy, and atomic force microscopy, the spherical titania nanostructures are identified to be anatase crystalline nanodisks with an average diameter of 9 nm and height of 0.5 nm. The nanodisks have a large Brunauer−Emmett−Teller specific surface area of 227 m2/g. The FT-IR and X-ray photoemission spectra of the nanodisks confirm that the skeleton structure of the titania nanodisk is formed through H-bonding of the −Ti−O−Ti− network through tetrahedrally coordinated vacancies designated 4Ti4+−OH. Analysis of the UV−vis and PL spectra reveals that the band-gap energy is red-shifted to 3.02 eV from that of TiO2 nanoparticle dots and its transition nature is exclusively indirect. The PL emission spectrum of the titania nanodisks exhibits a strong structural emission band around 420 nm with shoulders around 470 and 550 nm which is attributed to the transition from three different exciton-trapped surface states. In addition, another surface emission originating from the coordinatively unsaturated ions (Ti3+) is observed at 618 nm. These results suggest that coupling of the surface charge carriers with the lattice phonon of the nanostructures is so strong that the dominant route to charge recombination in titania nanodisks is nonradiative. Supporting the steady-state spectral observations, the decay profiles of the surface emission measured by using a femtosecond laser time-resolved PL system fit into a triexponential function with relatively longer lifetimes (20−30 ps, 1.1−1.5 ns, and 4.5−6.0 ns) as compared to those of simple nanoparticle dots, indicating that recombination of the charge carriers on the nanodisk surface is very prolonged. Being consistent with this, the photocatalytic efficiency for the reduction of methyl orange is much higher in the presence of the titania nanodisks than that observed in the presence of Degussa P-25.

Sum frequency generation (SFG) spectroscopic techniques are used to investigate the molecular orientation of adsorbed acetonitrile on rutile TiO2 (110) at the solid-vapor interface. Generally, most molecular orientation analyses using SFG have been performed on dielectric substrates, to avoid the spectral interference between resonant and the near-resonant background signal. Although rutile crystal can be treated as a dielectric substrate, its electronic state contributes to the intensity and interferes with the resonant signal when the SFG frequency is close to its band gap energy. In addition, the rutile crystal is a uniaxial birefringent material, and the (110) surface is anisotropic, which further complicates the spectral analysis. In this study, various SFG measurement techniques were applied, and quantitative analytical methods were established to interpret the surface orientation of an adsorbed molecule. SFG vibrational spectra of acetonitrile on rutile TiO2 (110) surface have been measured using distinct polarization combinations, polarization mapping, and null angle method. By varying the polarization combinations of SFG, the magnitude and shape of the spectra undergo substantial change, which originate from the interference between the near-resonant signal from the rutile substrate and the resonance signal from the acetonitrile. Theory, simulation, and analytical methods for obtaining quantitative orientation information of a molecule on an anisotropic semiconductor substrate in the presence of a near-resonant signal are presented.

The immunological synapse (IS) is one of the most pivotal communication strategies in immune cells. Understanding the molecular basis of the IS provides critical information regarding how immune cells mount an effective immune response. Fluorescence microscopy provides a fundamental tool to study the IS. However, current imaging techniques for studying the IS cannot sufficiently achieve high resolution in real cell-cell conjugates. Here we present a new device that allows for high-resolution imaging of the IS with conventional confocal microscopy in a high-throughput manner. Combining micropits and single cell trap arrays, we have developed a new microfluidic platform that allows visualization of the IS in vertically “stacked” cells. Using this vertical cell pairing (VCP) system, we investigated the dynamics of the inhibitory synapse mediated by an inhibitory receptor, programed death protein-1 (PD-1) and the cytotoxic synapse at the single cell level. In addition to the technique innovation, we demonstrated novel biological findings by this VCP device, including novel distribution of F-actin and cytolytic granules at the IS, PD-1 microclusters in the NK IS, and kinetics of cytotoxicity. We propose that this high-throughput, cost-effective, easy-to-use VCP system, along with conventional imaging techniques, can be used to address a number of significant biological questions in a variety of disciplines.