Extended depth-of-field microscopy with a high-speed deformable mirror.

We present a wide-field fluorescence microscopy add-on that provides a fast, light-efficient extended depth-of-field (EDOF) using a deformable mirror with an update rate of 20 kHz. Out-of-focus contributions in the raw EDOF images are suppressed with a deconvolution algorithm derived directly from the microscope 3D optical transfer function. Demonstrations of the benefits of EDOF microscopy are shown with GCaMP-labeled mouse brain tissue.

Band Gap Engineering with Ultralarge Biaxial Strains in Suspended Monolayer MoS2.

We demonstrate the continuous and reversible tuning of the optical band gap of suspended monolayer MoS2 membranes by as much as 500 meV by applying very large biaxial strains. By using chemical vapor deposition (CVD) to grow crystals that are highly impermeable to gas, we are able to apply a pressure difference across suspended membranes to induce biaxial strains. We observe the effect of strain on the energy and intensity of the peaks in the photoluminescence (PL) spectrum and find a linear tuning rate of the optical band gap of 99 meV/%. This method is then used to study the PL spectra of bilayer and trilayer devices under strain and to find the shift rates and Grüneisen parameters of two Raman modes in monolayer MoS2. Finally, we use this result to show that we can apply biaxial strains as large as 5.6% across micron-sized areas and report evidence for the strain tuning of higher level optical transitions.

Dictionary-based image reconstruction for superresolution in integrated circuit imaging.

Resolution improvement through signal processing techniques for integrated circuit imaging is becoming more crucial as the rapid decrease in integrated circuit dimensions continues. Although there is a significant effort to push the limits of optical resolution for backside fault analysis through the use of solid immersion lenses, higher order laser beams, and beam apodization, signal processing techniques are required for additional improvement. In this work, we propose a sparse image reconstruction framework which couples overcomplete dictionary-based representation with a physics-based forward model to improve resolution and localization accuracy in high numerical aperture confocal microscopy systems for backside optical integrated circuit analysis. The effectiveness of the framework is demonstrated on experimental data.

Effect of vector asymmetry of radially polarized beams in solid immersion microscopy.

We theoretically and experimentally investigate the effect of imperfect vector symmetry on radially polarized beams focused by an aplanatic solid immersion lens at a numerical aperture of 3.3. We experimentally achieve circularly symmetric focused spot with a full-width-half-maximum of ~λ0/5.7 at λ0 = 1,310 nm, free-space wavelength. The tight spatial confinement and overall circular symmetry of the focused radially polarized beam are found to be sensitive to perturbations of its cylindrical polarization symmetry. The addition of a liquid crystal based variable retarder to the optical path can effectively ensure the vector symmetry and achieve circularly symmetric focused spots at such high numerical aperture conditions.

Evanescent waves in high numerical aperture aplanatic solid immersion microscopy: effects of forbidden light on subsurface imaging.

The collection of light at very high numerical aperture allows detection of evanescent waves above the critical angle of total internal reflection in solid immersion lens microscopy. We investigate the effect of such evanescent modes, so-called forbidden light, on the far-field imaging properties of an aplanatic solid immersion microscope by developing a dyadic Green's function formalism in the context of subsurface semiconductor integrated circuit imaging. We demonstrate that the collection of forbidden light allows for sub-diffraction spatial resolution and substantial enhancement of photon collection efficiency albeit inducing wave-front discontinuities and aberrations.

Charge tuning of nonresonant magnetoexciton phonon interactions in graphene.

Far from resonance, the coupling of the G-band phonon to magnetoexcitons in single layer graphene displays kinks and splittings versus filling factor that are well described by Pauli blocking and unblocking of inter- and intra-Landau level transitions. We explore the nonresonant electron-phonon coupling by high-magnetic field Raman scattering while electrostatic tuning of the carrier density controls the filling factor. We show qualitative and quantitative agreement between spectra and a linearized model of electron-phonon interactions in magnetic fields. The splitting is caused by dichroism of left- and right-handed circular polarized light due to lifting of the G-band phonon degeneracy, and the piecewise linear slopes are caused by the linear occupancy of sequential Landau levels versus ν.

How graphene slides: measurement and theory of strain-dependent frictional forces between graphene and SiO2.

Strain, bending rigidity, and adhesion are interwoven in determining how graphene responds when pulled across a substrate. Using Raman spectroscopy of circular, graphene-sealed microchambers under variable external pressure, we demonstrate that graphene is not firmly anchored to the substrate when pulled. Instead, as the suspended graphene is pushed into the chamber under pressure, the supported graphene outside the microchamber is stretched and slides, pulling in an annulus. Analyzing Raman G band line scans with a continuum model extended to include sliding, we extract the pressure dependent sliding friction between the SiO2 substrate and mono-, bi-, and trilayer graphene. The sliding friction for trilayer graphene is directly proportional to the applied load, but the friction for monolayer and bilayer graphene is inversely proportional to the strain in the graphene, which is in violation of Amontons' law. We attribute this behavior to the high surface conformation enabled by the low bending rigidity and strong adhesion of few layer graphene.

A national professional development program fills mentoring gaps for postdoctoral researchers.

The Postdoc Academy: Succeeding as a Postdoc was designed to build postdocs' skills in career transition, career planning, collaborative research, resilience, and self-reflection. This study examined self-reported changes in five skills as learners progressed through the course. Data were collected from participants who responded to both pre- and post-surveys and engaged with the course learning activities. Results from repeated measures multivariate analysis of variance revealed that all of the self-reported perceptions of skills improved significantly upon completion of the course. Hierarchical regressions revealed that underrepresented minority learners had greater gains in their development of skills in career planning, resilience, and self-reflection. Qualitative analysis of learners' responses to learning activities found that postdocs perceived networking and mentor support as contributing factors to their skill advancement while tensions among multiple obligations and concerns of uncertainties were significant challenges to applying those skills.

Training in the art and science of facilitation to scale research mentor training in low and middle income countries.

Advancing biomedical research in low and middle income countries (LMICs) to expand the capacity for LMICs to integrate biomedical research into their health care systems and education has been the focus of many programs in global health over the past two decades. Central to the success of these programs is effective research mentoring, characterized by academic, career and psychosocial support through culturally appropriate practices. Research mentoring is a learned skill, developed through training, mutual discussions, practice and feedback. The majority of extant training programs are designed and delivered by US partners, so the next stage in building capacity is to train facilitators within the LMIC partner institutions to contextualize and advance mentoring specifically within their cultural and institutional norms by co-developing, delivering and evaluating semi-annual research mentoring training. To this end, we describe the development, delivery and outcome evaluation of a 5-week course in the art and skill of facilitation. Care was taken to explicitly distinguish between concepts of "teaching" and "facilitation," since "teaching" is closely connected to a transmission or banking model of education, which is characterized by "top-down," hierarchical relationship. The course discussed power and positionality, themes that resonate with partners in Nigeria and Tanzania. These themes provided unique entry into deeper conversations core to advancing mentoring practice away from the traditional dyadic power structure that remains from colonization. Evaluation findings indicate significant advances in awareness of differences between teaching and facilitating, increased confidence in facilitation skills, especially in the area of structured planning and organization, as well as improved communication and interpersonal skills. All respondents felt that students in Nigeria and Tanzania would respond well to the facilitation approach conveyed during the course and they found value in participating in the course as a cohort.

Preparing future STEM faculty through flexible teaching professional development.

We have prepared thousands of future STEM faculty around the world to adopt evidence-based instructional practices through their participation in two massive open online courses (MOOCs) and facilitated in-person learning communities. Our novel combination of asynchronous online and coordinated, structured face-to-face learning community experiences provides flexible options for STEM graduate students and postdoctoral fellows to pursue teaching professional development. A total of 14,977 participants enrolled in seven offerings of the introductory course held 2014-2018, with 1,725 participants (11.5% of enrolled) completing the course. Our results of high levels of engagement and learning suggest that leveraging the affordances of educational technologies and the geographically clustered nature of this learner demographic in combination with online flexible learning could be a sustainable model for large scale professional development in higher education. The preparation of future STEM faculty makes an important difference in establishing high-quality instruction that meets the diverse needs of all undergraduate students, and the initiative described here can serve as a model for increasing access to such preparation.

Label-free and dynamic detection of biomolecular interactions for high-throughput microarray applications.

Direct monitoring of primary molecular-binding interactions without the need for secondary reactants would markedly simplify and expand applications of high-throughput label-free detection methods. A simple interferometric technique is presented that monitors the optical phase difference resulting from accumulated biomolecular mass. As an example, 50 spots for each of four proteins consisting of BSA, human serum albumin, rabbit IgG, and protein G were dynamically monitored as they captured corresponding antibodies. Dynamic measurements were made at 26 pg/mm(2) SD per spot and with a detectable concentration of 19 ng/ml. The presented method is particularly relevant for protein microarray analysis because it is label-free, simple, sensitive, and easily scales to high-throughput.

Biaxial strain in graphene adhered to shallow depressions.

Measurements on graphene exfoliated over a substrate prepatterned with shallow depressions demonstrate that graphene does not remain free-standing but instead adheres to the substrate despite the induced biaxial strain. The strain is homogeneous over the depression bottom as determined by Raman measurements. We find higher Raman shifts and Gruneisen parameters of the phonons underlying the G and 2D bands under biaxial strain than previously reported. Interference modeling is used to determine the vertical position of the graphene and to calculate the optimum dielectric substrate stack for maximum Raman signal.

Theory of change models deepen online learning evaluation.

Massive Open Online Courses (MOOCs) have gained traction as resources for professional development. This article presents the method that we used to evaluate a professional development MOOC for postdoctoral trainees that was created by a university consortium in the US. Most approaches to evaluating MOOCs focus on analysis of participation, outcomes from course assignments, self-reported learning outcomes, course completion and user pathways through the online content or clickstream data. Few published evaluations describe in detail how learning happens within online courses and the anticipated medium and longer term cognitive and behavioral outcomes on participants. This work aims to guide those who are designing, implementing and evaluating MOOCs through applying theories of change to focus evaluation on the process of learning. This approach can be used as a complement to traditional approaches for evaluating MOOCs. We described how we worked with the MOOC team building the content to develop a theory of change for each module (or lesson) within the MOOC and used the theory of change to guide evaluation of short and medium term participant cognitive, affective and attitudinal, and behavioral outcomes. Finally, we share lessons learned and suggestions for implementing theories of change in both the design and evaluation phases of MOOC development.

Direct observation of conformation of a polymeric coating with implications in microarray applications.

The conformation of a three-dimensional polymeric coating (copoly(DMA-NAS-MAPS)) and immobilization and hybridization of DNA strands on the polymer coated surface are investigated. A conformational change, specifically the swelling of the surface adsorbed polymer upon hydration, is quantified in conjunction with the application of this polymer coating for DNA microarray applications. Fluorescently labeled short DNA strands (23mers) covalently linked to the functional groups on the adsorbed polymer are used as probes to measure the swelling of the polymer. A fluorescence microscopy technique, Spectral Self-Interference Fluorescence Microscopy (SSFM), is utilized to directly measure the change in axial position of fluorophores due to swelling with subnanometer accuracy. Additionally, immobilization characteristics of single stranded DNA (ssDNA) and double stranded DNA (dsDNA) probes, as well as hybridization of ssDNA with target strands have been studied. The results show that ssDNA further away from the surface is hybridized more efficiently, which strengthens the earlier analysis of this polymeric coating as a simple but highly efficient and robust DNA microarray surface chemistry.

Widefield subsurface microscopy of integrated circuits.

We apply the numerical aperture increasing lens technique to widefield subsurface imaging of silicon integrated circuits. We demonstrate lateral and longitudinal resolutions well beyond the limits of conventional backside imaging. With a simple infrared widefield microscope (lambda(0) = 1.2 microm), we demonstrate a lateral spatial resolution of 0.26 microm (0.22 lambda(0)) and a longitudinal resolution of 1.24 microm (1.03 lambda(0)) for backside imaging through the silicon substrate of an integrated circuit. We present a spatial resolution comparison between widefield and confocal microscopy, which are essential in integrated circuit analysis for emission and excitation microscopy, respectively.

Resonant Cavity Imaging: A Means Toward High-Throughput Label-Free Protein Detection.

The resonant cavity imaging biosensor (RCIB) is an optical technique for detecting molecular binding interactions label free at many locations in parallel that employs an optical resonant cavity for high sensitivity. Near-infrared light centered at 1512.5 nm couples resonantly through a Fabry-Perot cavity constructed from dielectric reflectors (Si/SiO(2)), one of which serves as the binding surface. As the wavelength is swept using a tunable laser, a near-infrared digital camera monitors cavity transmittance at each pixel. A wavelength shift in the local resonant response of the optical cavity indicates binding. Positioning the sensing surface with respect to the standing wave pattern of the electric field within the cavity controls the sensitivity with which the presence of bound molecules is detected. Transmitted intensity at thousands of pixel locations is recorded simultaneously in a 10 s, 5 nm scan. An initial proof-of-principle setup has been constructed. A test sample was fabricated with 25, 100-mum wide square features, each with a different density of 1-mum square depressions etched 12 nm into the SiO(2) surface. The average depth of each etched region was found with 0.05 nm rms precision. In a second test, avidin, bound selectively to biotin conjugated bovine serum albumin, was detected.

Long tailed trions in monolayer MoS2: Temperature dependent asymmetry and resulting red-shift of trion photoluminescence spectra.

Monolayer molybdenum disulfide (MoS2) has emerged as a model system for studying many-body physics because the low dimensionality reduces screening leading to tightly bound states stable at room temperature. Further, the many-body states possess a pseudo-spin degree of freedom that corresponds with the two direct-gap valleys of the band structure, which can be optically manipulated. Here we focus on one bound state, the negatively charged trion. Unlike excitons, trions can radiatively decay with non-zero momentum by kicking out an electron, resulting in an asymmetric trion photoluminescence (PL) peak with a long low-energy tail and peak position that differs from the zero momentum trion energy. The asymmetry of the trion PL peak and resulting peak red-shift depends both on the trion size and a temperature-dependent contribution. Ignoring the trion asymmetry will result in over estimating the trion binding energy by nearly 20 meV at room temperature. We analyze the temperature-dependent PL to reveal the effective trion size, consistent with the literature, and the temperature dependence of the band gap and spin-orbit splitting of the valence band. This is the first time the temperature-dependence of the trion PL has been analyzed with such detail in any system.

MRI of the lung gas-space at very low-field using hyperpolarized noble gases.

In hyperpolarized (HP) noble-gas magnetic resonance imaging, large nuclear spin polarizations, about 100,000 times that ordinarily obtainable at thermal equilibrium, are created in 3He and 129Xe. The enhanced signal that results can be employed in high-resolution MRI studies of void spaces such as in the lungs. In HP gas MRI the signal-to-noise ratio (SNR) depends only weakly on the static magnetic field (B(0)), making very low-field (VLF) MRI possible; indeed, it is possible to contemplate portable MRI using light-weight solenoids or permanent magnets. This article reports the first in vivo VLF MR images of the lungs in humans and in rats, obtained at a field of only 15 millitesla (150 Gauss).

Digital sensing and sizing of vesicular stomatitis virus pseudotypes in complex media: a model for Ebola and Marburg detection.

Rapid, sensitive, and direct label-free capture and characterization of nanoparticles from complex media such as blood or serum will broadly impact medicine and the life sciences. We demonstrate identification of virus particles in complex samples for replication-competent wild-type vesicular stomatitis virus (VSV), defective VSV, and Ebola- and Marburg-pseudotyped VSV with high sensitivity and specificity. Size discrimination of the imaged nanoparticles (virions) allows differentiation between modified viruses having different genome lengths and facilitates a reduction in the counting of nonspecifically bound particles to achieve a limit-of-detection (LOD) of 5 × 10(3) pfu/mL for the Ebola and Marburg VSV pseudotypes. We demonstrate the simultaneous detection of multiple viruses in a single sample (composed of serum or whole blood) for screening applications and uncompromised detection capabilities in samples contaminated with high levels of bacteria. By employing affinity-based capture, size discrimination, and a "digital" detection scheme to count single virus particles, we show that a robust and sensitive virus/nanoparticle sensing assay can be established for targets in complex samples. The nanoparticle microscopy system is termed the Single Particle Interferometric Reflectance Imaging Sensor (SP-IRIS) and is capable of high-throughput and rapid sizing of large numbers of biological nanoparticles on an antibody microarray for research and diagnostic applications.