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.

The extensive phenotypic and functional heterogeneity of cancer cells plays an important role in tumor progression and therapeutic resistance. Characterizing this heterogeneity and identifying invasive phenotype may provide possibility to improve chemotherapy treatment. By mimicking cancer cell perfusion through circulatory system in metastasis, we develop a unique microfluidic cytometry (MC) platform to separate cancer cells at high throughput, and further derive a physical parameter ‘transportability’ to characterize the ability to pass through micro-constrictions. The transportability is determined by cell stiffness and cell-surface frictional property, and can be used to probe tumor heterogeneity, discriminate more invasive phenotypes and correlate with biomarker expressions in breast cancer cells. Decreased cell stiffness and cell-surface frictional force leads to an increase in transportability and may be a feature of invasive cancer cells by promoting cell perfusion through narrow spaces in circulatory system. The MC-Chip provides a promising microfluidic platform for studying cell mechanics and transportability could be used as a novel marker for probing tumor heterogeneity and determining invasive phenotypes.

A new class of nanostructures were fabricated by one-step hydrothermal reaction of a mixture solution of TiO 2 anatase powder and a Sn-porphyrin, trans-dihydroxo [5,10,15,20-tetrakis(p-tolyl)porphyrinato]tin(IV) [SnTTP], and they were found to be well-crystalline trititanate (H 2 Ti 3 O 7 )-type multilayered nanofibers (TiNFs) intercalated by SnTTP which have lengths in the range of 0.5-1 µm with an average diameter of approximately 50 nm. Based on the femtosecond-diffuse reflectance transient absorption and photoluminescence spectroscopic measurements, the SnTTP-intercalated TiNFs were observed to exhibit efficient optoelectronic properties such as photoinduced electron transfer from deep surface states of trititanate layer to SnTTP, forming an anion radical SnTTP .•rapidly in a few picoseconds. These results infer that electrons and holes are effectively separated in the SnTTP-TiNFs upon illumination, and consequently remarkable UV-visible light-sensitive photocatalytic activities as compared to those of free TiNFs and SnTTP, suggesting that the SnTTP-intercalated TiNFs have potential application in development of efficient artificial photosynthetic systems and photoelectronic materials.

Monitoring and understanding monomer conformational changes as substituents are varied is an important step in investigating in situ polymerization and controlling the polymerization process as well as regulating the functionality of polymer surfaces to provide efficient polymer coatings. In this study, the preliminary stage involves characterization of the interfacial structures of methacrylate-based functional monomers and their polymer thin films at the air−liquid interface using femtosecond sum frequency generation spectroscopy (FSFGS). By varying the substituted ethyl group of the methacrylate monomers with hydroxy (−OH), chloro (−Cl), and phenoxy (−OPh), the alphamethyl (α-CH 3 ), alkene-methylene (alkene-CH 2 ), and methylene (CH 2 ) groups are observed to have preferential surface ordering toward the air interface. A peak positioned at ∼3000 cm −1 was observed in the spectrum of the 2-hydroxyethyl methacrylate (HEMA) monomer and assigned to alkene-CH 2 group. This functional group signifies that the pure monomer has not undergone polymerization. However, the SFG spectra of the polymer versions of these monomers revealed that for the poly(2hydroxyethyl methacrylate), the α-CH 3 group dominated the surface; in both poly(2-chloroethyl methacrylate) and poly(2phenoxyethyl methacrylate), the α-CH 3 and CH 2 groups were both segregated at the interface. These observations of the conformational changes of the monomer units and polymer thin films indicated that substitution of the ethyl group of the methacrylates affected the behavior of the interfacial molecules.

This study employs femtosecond sum frequency generation (FS-SFG) spectroscopy to identify the functional groups at the air–liquid interface of the 2-methoxyethyl methacrylate (MEMA) monomer. Based on the collected spectra, the methoxy group (−OCH3), the methylene (−CH2) group from the ethyl side chain of MEMA, the α-methyl group (α-CH3), and the alkene-methylene (CH2) groups are present at the interface. Tilt angle was determined by calculating the intensity ratio values from the fitting results, and then the ratios are compared to the simulated SFG curves. The results derived from the conventional polarization combination (SSP, PPP, and PSS) approach show average tilt angles of 60° for −OCH3 symmetric stretch vibration dipoles. Using the polarization mapping method, the tilt angle for the symmetric stretch of the −OCH3 group was 48° with an intensity ratio (SSP/PPP) of 17. The orientation distribution of the functional groups was also obtained from the amplitude ratios using the two methods. In conclusion, the MEMA monomer is partially ordered at the air–liquid interface as a result of influences from its different functional groups. These results suggest that the α-CH3 and −OCH3 symmetric stretches are tilted further away from the surface normal, thus deviating from the presumed model of well-ordered interfacial molecules.