Browsing by Author "Chen, Jixin"

Now showing 1 - 6 of 6
  • Thumbnail Image
    Item
    Charge-Dependent Transport Switching of Single Molecular Ions in a Weak Polyelectrolyte Multilayer
    (American Chemical Society, 2014) Tauzin, Lawrence J.; Shuang, Bo; Kisley, Lydia; Mansur, Andrea P.; Chen, Jixin; de Leon, Al; Advincula, Rigoberto C.; Landes, Christy F.
    The tunable nature of weak polyelectrolyte multilayers makes them ideal candidates for drug loading and delivery, water filtration, and separations, yet the lateral transport of charged molecules in these systems remains largely unexplored at the single molecule level. We report the direct measurement of the charge-dependent, pH-tunable, multimodal interaction of single charged molecules with a weak polyelectrolyte multilayer thin film, a 10 bilayer film of poly(acrylic acid) and poly(allylamine hydrochloride) PAA/PAH. Using fluorescence microscopy and single-molecule tracking, two modes of interaction were detected: (1) adsorption, characterized by the molecule remaining immobilized in a subresolution region and (2) diffusion trajectories characteristic of hopping (D ∼ 10–9 cm2/s). Radius of gyration evolution analysis and comparison with simulated trajectories confirmed the coexistence of the two transport modes in the same single molecule trajectories. A mechanistic explanation for the probe and condition mediated dynamics is proposed based on a combination of electrostatics and a reversible, pH-induced alteration of the nanoscopic structure of the film. Our results are in good agreement with ensemble studies conducted on similar films, confirm a previously-unobserved hopping mechanism for charged molecules in polyelectrolyte multilayers, and demonstrate that single molecule spectroscopy can offer mechanistic insight into the role of electrostatics and nanoscale tunability of transport in weak polyelectrolyte multilayers.
  • Thumbnail Image
    Item
    Fast Step Transition and State Identification (STaSI) for Discrete Single-Molecule Data Analysis
    (American Chemical Society, 2014) Shuang, Bo; Cooper, David; Taylor, J. Nick; Kisley, Lydia; Chen, Jixin; Wang, Wenxiao; Li, Chun Biu; Komatsuzaki, Tamiki; Landes, Christy F.; Rice Quantum Institute
    We introduce a step transition and state identification (STaSI) method for piecewise constant single-molecule data with a newly derived minimum description length equation as the objective function. We detect the step transitions using the Student’s t test and group the segments into states by hierarchical clustering. The optimum number of states is determined based on the minimum description length equation. This method provides comprehensive, objective analysis of multiple traces requiring few user inputs about the underlying physical models and is faster and more precise in determining the number of states than established and cutting-edge methods for single-molecule data analysis. Perhaps most importantly, the method does not require either time-tagged photon counting or photon counting in general and thus can be applied to a broad range of experimental setups and analytes.
  • Thumbnail Image
    Item
    High ionic strength narrows the population of sites participating in protein ion-exchange adsorption: A single-molecule study
    (Elsevier, 2014) Kisley, Lydia; Chen, Jixin; Mansur, Andrea P.; Dominguez-Medina, Sergio; Kulla, Eliona; Kang, Marci; Shuang, Bo; Kourentzi, Katerina; Poongavanam, Mohan-Vivekanandan; Dhamane, Sagar; Willson, Richard C.; Landes, Christy F.
    The retention and elution of proteins in ion-exchange chromatography is routinely controlled by adjusting the mobile phase salt concentration. It has repeatedly been observed, as judged from adsorption isotherms, that the apparent heterogeneity of adsorption is lower at more-eluting, higher ionic strength. Here, we present an investigation into the mechanism of this phenomenon using a single-molecule, super-resolution imaging technique called motion-blur Points Accumulation for Imaging in Nanoscale Topography (mbPAINT). We observed that the number of functional adsorption sites was smaller at high ionic strength and that these sites had reduced desorption kinetic heterogeneity, and thus narrower predicted elution profiles, for the anion-exchange adsorption of ?-lactalbumin on an agarose-supported, clustered-charge ligand stationary phase. Explanations for the narrowing of the functional population such as inter-protein interactions and protein or support structural changes were investigated through kinetic analysis, circular dichroism spectroscopy, and microscopy of agarose microbeads, respectively. The results suggest the reduction of heterogeneity is due to both electrostatic screening between the protein and ligand and tuning the steric availability within the agarose support. Overall, we have shown that single molecule spectroscopy can aid in understanding the influence of ionic strength on the population of functional adsorbent sites participating in the ion-exchange chromatographic separation of proteins.
  • Thumbnail Image
    Item
    Single-Molecule FRET Studies of HIV TAR–DNA Hairpin Unfolding Dynamics
    (American Chemical Society, 2014) Chen, Jixin; Poddar, Nitesh K.; Tauzin, Lawrence J.; Cooper, David; Kolomeisky, Anatoly B.; Landes, Christy F.
    We directly measure the dynamics of the HIV trans-activation response (TAR)-DNA hairpin with multiple loops using single-molecule Förster resonance energy transfer (smFRET) methods. Multiple FRET states are identified that correspond to intermediate melting states of the hairpin. The stability of each intermediate state is calculated from the smFRET data. The results indicate that hairpin unfolding obeys a "fraying and peeling" mechanism, and evidence for the collapse of the ends of the hairpin during folding is observed. These results suggest a possible biological function for hairpin loops serving as additional fraying centers to increase unfolding rates in otherwise stable systems. The experimental and analytical approaches developed in this article provide useful tools for studying the mechanism of multistate DNA hairpin dynamics and of other general systems with multiple parallel pathways of chemical reactions.
  • Thumbnail Image
    Item
    Super-Resolution mbPAINT for Optical Localization of Single-Stranded DNA
    (American Chemical Society, 2013) Chen, Jixin; Bremauntz, Alberto; Kisley, Lydia; Shuang, Bo; Landes, Christy F.
    We demonstrate the application of superlocalization microscopy to identify sequence-specific portions of single-stranded DNA (ssDNA) with sequence resolution of 50 nucleotides, corresponding to a spatial resolution of 30 nm. Super-resolution imaging was achieved using a variation of a single-molecule localization method, termed as ﾓmotion blurﾔ point accumulation for imaging in nanoscale topography (mbPAINT). The target ssDNA molecules were immobilized on the substrate. Short, dye-labeled, and complementary ssDNA molecules stochastically bound to the target ssDNA, with repeated binding events allowing super-resolution. Sequence specificity was demonstrated via the use of a control, noncomplementary probe. The results support the possibility of employing relatively inexpensive short ssDNAs to identify gene sequence specificity with improved resolution in comparison to the existing methods.
  • Thumbnail Image
    Item
    Troika of single particle tracking programing: SNR enhancement, particle identification, and mapping
    (Royal Society of Chemistry, 2014) Shuang, Bo; Chen, Jixin; Kisley, Lydia; Landes, Christy F.
    Single particle tracking (SPT) techniques provide a microscopic approach to probe in vivo and in vitro structure and reactions. Automatic analysis of SPT data with high efficiency and accuracy spurs the development of SPT algorithms. In this perspective, we review a range of available techniques used in SPT analysis programs. In addition, we present an example SPT program step-by-step to provide a guide so that researchers can use, modify, and/or write a SPT program for their own purposes.