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Transition metal dichalcogenides (TMDs) are regarded as a possible material platform for quantum information science and related device applications. In TMD monolayers, the dephasing time and inhomogeneity are crucial parameters for any quantum information application. In TMD heterostructures, coupling strength and interlayer exciton lifetimes are also parameters of interest. However, many demonstrations in TMDs can only be realized at specific spots on the sample, presenting a challenge to the scalability of these applications. Here, using multi-dimensional coherent imaging spectroscopy, we shed light on the underlying physics—including dephasing, inhomogeneity, and strain—for a MoSe2 monolayer and identify both promising and unfavorable areas for quantum information applications. We, furthermore, apply the same technique to a MoSe2/WSe2 heterostructure. Despite the notable presence of strain and dielectric environment changes, coherent and incoherent coupling and interlayer exciton lifetimes are mostly robust across the sample. This uniformity is despite a significantly inhomogeneous interlayer exciton photoluminescence distribution that suggests a bad sample for device applications. This robustness strengthens the case for TMDs as a next-generation material platform in quantum information science and beyond. [ABSTRACT FROM AUTHOR]

Epigenetic mechanisms regulate the multilineage differentiation capacity of hematopoietic stem cells (HSCs) into a variety of blood and immune cells. Mapping the chromatin dynamics of functionally defined cell populations will shed mechanistic insight into 2 major, unanswered questions in stem cell biology: how does epigenetic identity contribute to a cell type's lineage potential, and how do cascades of chromatin remodeling dictate ensuing fate decisions? Our recent work revealed evidence of multilineage gene priming in HSCs, where open cis- regulatory elements (CREs) exclusively shared between HSCs and unipotent lineage cells were enriched for DNA binding motifs of known lineage-specific transcription factors. Oligopotent progenitor populations operating between the HSCs and unipotent cells play essential roles in effecting hematopoietic homeostasis. To test the hypothesis that selective HSC-primed lineage-specific CREs remain accessible throughout differentiation, we used ATAC-seq to map the temporal dynamics of chromatin remodeling during progenitor differentiation. We observed epigenetic-driven clustering of oligopotent and unipotent progenitors into distinct erythromyeloid and lymphoid branches, with multipotent HSCs and MPPs associating with the erythromyeloid lineage. We mapped the dynamics of lineage-primed CREs throughout hematopoiesis and identified both unique and shared CREs as potential lineage reinforcement mechanisms at fate branch points. Additionally, quantification of genome-wide peak count and size revealed overall greater chromatin accessibility in HSCs, allowing us to identify HSC-unique peaks as putative regulators of self-renewal and multilineage potential. Finally, CRISPRi-mediated targeting of ATACseq-identified putative CREs in HSCs allowed us to demonstrate the functional role of selective CREs in lineage-specific gene expression. These findings provide insight into the regulation of stem cell multipotency and lineage commitment throughout hematopoiesis and serve as a resource to test functional drivers of hematopoietic lineage fate. [ABSTRACT FROM AUTHOR]

A growing body of evidence suggests that L-selectin ligands presented on circulating tumor cells facilitate metastasis by binding L-selectin presented on leukocytes. Commonly used methods for detecting L-selectin ligands on tissues, e.g., immunostaining, are performed under static, no-flow conditions. However, such analysis does not assay for functional L-selectin ligands, specifically those ligands that promote adhesion under shear flow conditions. Recently our lab developed a method, termed dynamic biochemical tissue analysis (DBTA), to detect functional selectin ligands in situ by probing tissues with L-selectin-coated microspheres under hemodynamic flow conditions. In this investigation, DBTA was used to probe human colon tissues for L-selectin ligand activity. The detection of L-selectin ligands using DBTA was highly specific. Furthermore, DBTA reproducibly detected functional L-selectin ligands on diseased, e.g., cancerous or inflamed, tissues but not on noncancerous tissues. In addition, DBTA revealed a heterogeneous distribution of functional L-selectin ligands on colon cancer tissues. Most notably, detection of L-selectin ligands by immunostaining using HECA-452 antibody only partially correlated with functional L-selectin ligands detected by DBTA. In summation, the results of this study demonstrate that DBTA detects functional selectin ligands to provide a unique characterization of pathological tissue. [ABSTRACT FROM AUTHOR]

We report ultrabroadband two-dimensional electronic spectroscopy (2D ES) measurements obtained in the pump-probe geometry using conventional optics. A phase-stabilized Michelson interferometer provides the pump-pulse delay interval, τ 1 , necessary to obtain the excitation-frequency dimension. Spectral resolution of the probe beam provides the detection-frequency dimension, ω 3 . The interferometer incorporates active phase stabilization via a piezo stage and feedback from interference of a continuous-wave reference laser detected in quadrature. To demonstrate the method, we measured a well-characterized laser dye sample and obtained the known peak structure. The vibronic peaks are modulated as a function of the waiting time, τ 2 , by vibrational wave packets. The interferometer simplifies ultrabroadband 2D ES measurements and analysis.

We demonstrate rapid imaging based on four-wave mixing (FWM) by assessing the quality of advanced materials through measurement of their nonlinear response, exciton dephasing, and exciton lifetimes. We use a WSe 2 monolayer grown by chemical vapor deposition as a canonical example to demonstrate these capabilities. By comparison, we show that extracting material parameters such as FWM intensity, dephasing times, excited state lifetimes, and distribution of dark/localized states allows for a more accurate assessment of the quality of a sample than current prevalent techniques, including white light microscopy and linear micro-reflectance spectroscopy. We further discuss future improvements of the ultrafast FWM techniques by modeling the robustness of exponential decay fits to different spacing of the sampling points. Employing ultrafast nonlinear imaging in real-time at room temperature bears the potential for rapid in-situ sample characterization of advanced materials and beyond.

We show that accelerated nonlinear imaging, such as stimulated Raman scattering and pump-probe imaging, is enabled by an order of magnitude reduction of data acquisition time when replacing the exponentially-weighted-moving-average low-pass filter in a lock-in amplifier with a simple-moving-average filter. We show that this simple-moving-average (box) lock-in yields a superior signal-to-noise ratio and suppression of extraneous modulations with short pixel dwell times, if one condition for the relation between the lock-in time constant and modulation frequencies is met. Our results, both theoretical and experimental, indicate that for nonlinear imaging applications, the box lock-in significantly outperforms conventional lock-in detection. These results facilitate the application of ultrafast and nonlinear imaging as a new standard for material characterization.

Hematopoiesis is arguably one of the best understood stem cell systems; however, significant challenges remain to reach a consensus understanding of the lineage potential, heterogeneity, and relationships of hematopoietic stem and progenitor cell populations. To gain new insights, we performed quantitative analyses of mature cell production from hematopoietic stem cells (HSCs) and multiple hematopoietic progenitor populations. Assessment of the absolute numbers of mature cell types produced by each progenitor cell revealed a striking erythroid dominance of all myeloid-competent progenitors assessed, accompanied by strong platelet reconstitution. All populations with myeloid potential also produced robust numbers of red blood cells and platelets in vivo. Clonal analysis by single-cell transplantation and by spleen colony assays revealed that a significant fraction of HSCs and multipotent progenitors have multilineage potential at the single-cell level. These new insights prompt an erythroid-focused model of hematopoietic differentiation., (Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.)

Epigenetic regulation serves as the basis for stem cell differentiation into distinct cell types, but it is unclear how global epigenetic changes are regulated during this process. Here, we tested the hypothesis that global chromatin organization affects the lineage potential of stem cells and that manipulation of chromatin dynamics influences stem cell function. Using nuclease sensitivity assays, we found a progressive decrease in chromatin digestion among pluripotent embryonic stem cells (ESCs), multipotent hematopoietic stem cells (HSCs), and mature hematopoietic cells. Quantitative high-resolution microscopy revealed that ESCs contain significantly more euchromatin than HSCs, with a further reduction in mature cells. Increased cellular maturation also led to heterochromatin localization to the nuclear periphery. Functionally, prevention of heterochromatin formation by inhibition of the histone methyltransferase G9A resulted in delayed HSC differentiation. Our results demonstrate global chromatin rearrangements during stem cell differentiation and that heterochromatin formation by H3K9 methylation regulates HSC differentiation., (Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.)

Laboratory scale to industrial scale purification of biomolecules from cell culture supernatants and lysed cell solutions can be accomplished using affinity chromatography. While affinity chromatography using porous protein A agarose beads packed in columns is arguably the most common method of laboratory scale isolation of antibodies and recombinant proteins expressing Fc fragments of IgG, it can be a time consuming and expensive process. Time and financial constraints are especially daunting in small basic science labs that must recover hundreds of micrograms to milligram quantities of protein from dilute solutions, yet lack access to high pressure liquid delivery systems and/or personnel with expertise in bioseparations. Moreover, product quantification and characterization may also excessively lengthen processing time over several workdays and inflate expenses (consumables, wages, etc.). Therefore, a fast, inexpensive, yet effective protocol is needed for laboratory scale isolation and characterization of antibodies and other proteins possessing an Fc fragment. To this end, we have devised a protocol that can be completed by limited-experience technical staff in less than 9 hr (roughly one workday) and as quickly as 4 hr, as opposed to traditional methods that demand 20+ work hours. Most required equipment is readily available in standard biomedical science, biochemistry, and (bio)chemical engineering labs, and all reagents are commercially available. To demonstrate this protocol, representative results are presented in which chimeric murine galectin-1 fused to human Fc (Gal-1hFc) from cell culture supernatant was isolated using a protein A membrane adsorber. Purified Gal-1hFc was quantified using an expedited Western blotting analysis procedure and characterized using flow cytometry. The streamlined workflow can be modified for other Fc-expressing proteins, such as antibodies, and/or altered to incorporate alternative quantification and characterization methods.

Multi-color immunofluorescence microscopy to detect specific molecules in the cell membrane can be coupled with parallel plate flow chamber assays to investigate mechanisms governing cell adhesion under dynamic flow conditions. For instance, cancer cells labeled with multiple fluorophores can be perfused over a potentially reactive substrate to model mechanisms of cancer metastasis. However, multi-channel single camera systems and color cameras exhibit shortcomings in image acquisition for real-time live cell analysis. To overcome these limitations, we used a dual camera emission splitting system to simultaneously capture real-time image sequences of fluorescently labeled cells in the flow chamber. Dual camera emission splitting systems filter defined wavelength ranges into two monochrome CCD cameras, thereby simultaneously capturing two spatially identical but fluorophore-specific images. Subsequently, psuedocolored one-channel images are combined into a single real-time merged sequence that can reveal multiple target molecules on cells moving rapidly across a region of interest.