A neural circuit state change underlying skilled movements.

In motor neuroscience, state changes are hypothesized to time-lock neural assemblies coordinating complex movements, but evidence for this remains slender. We tested whether a discrete change from more autonomous to coherent spiking underlies skilled movement by imaging cerebellar Purkinje neuron complex spikes in mice making targeted forelimb-reaches. As mice learned the task, millimeter-scale spatiotemporally coherent spiking emerged ipsilateral to the reaching forelimb, and consistent neural synchronization became predictive of kinematic stereotypy. Before reach onset, spiking switched from more disordered to internally time-locked concerted spiking and silence. Optogenetic manipulations of cerebellar feedback to the inferior olive bi-directionally modulated neural synchronization and reaching direction. A simple model explained the reorganization of spiking during reaching as reflecting a discrete bifurcation in olivary network dynamics. These findings argue that to prepare learned movements, olivo-cerebellar circuits enter a self-regulated, synchronized state promoting motor coordination. State changes facilitating behavioral transitions may generalize across neural systems.

The Paracrine Function of Mesenchymal Stem Cells in Response to Pulsed Focused Ultrasound.

We studied the paracrine function of mesenchymal stem cells (MSCs) derived from various sources in response to pulsed focused ultrasound (pFUS). Human adipose tissue (AD), bone marrow (BM), and umbilical cord (UC) derived MSCs were exposed to pFUS at two intensities: 0.45 W/cm2 ISATA (310 kPa PNP) and 1.3 W/cm2 ISATA (540 kPa PNP). Following pFUS, the viability and proliferation of MSCs were assessed using a hemocytometer and confocal microscopy, and their secreted cytokine profile determined using a multiplex ELISA. Our findings showed that pFUS can stimulate the production of immunomodulatory, anti-inflammatory, and angiogenic cytokines from MSCs which was dependent on both the source of MSC being studied and the acoustic intensity employed. These important findings set the foundation for additional mechanistic and validation studies using this novel noninvasive and clinically translatable technology for modulating MSC biology.

Density Based Characterization of Mechanical Cues on Cancer Cells Using Magnetic Levitation.

Extracellular matrix (ECM) stiffness is correlated to malignancy and invasiveness of cancer cells. Although the mechanism of change is unclear, mechanical signals from the ECM may affect physical properties of cells such as their density profile. The current methods, such as Percoll density-gradient centrifugation, are unable to detect minute density differences. A magnetic levitation device is developed (i.e., MagDense platform) where cells are levitated in a magnetic gradient allowing them to equilibrate to a levitation height that corresponds to their unique cellular density. In application of this system, MDA-MB-231 breast and A549 lung cancer cells are cultured and overall differences in cell density are observed in response to increasing collagen fiber density. Overall, density values are significantly more spread out for MDA-MB-231 cells extracted from the 1.44 mg mL-1 collagen gels compared to those from 0.72 mg mL-1 collagen, whereas no significant difference with A549 cell lines is observed. The MagDense platform can determine differences in cellular densities under various microenvironmental conditions. The imaging of cancer cells in a magnetic levitation device serves as a unique tool to observe changes in phenotypic properties of cancer cells as they relate to micromechanical cues encoded by the ECM.

Fast, in vivo voltage imaging using a red fluorescent indicator.

Genetically encoded voltage indicators (GEVIs) are emerging optical tools for acquiring brain-wide cell-type-specific functional data at unparalleled temporal resolution. To broaden the application of GEVIs in high-speed multispectral imaging, we used a high-throughput strategy to develop voltage-activated red neuronal activity monitor (VARNAM), a fusion of the fast Acetabularia opsin and the bright red fluorophore mRuby3. Imageable under the modest illumination intensities required by bright green probes (<50 mW mm-2), VARNAM is readily usable in vivo. VARNAM can be combined with blue-shifted optical tools to enable cell-type-specific all-optical electrophysiology and dual-color spike imaging in acute brain slices and live Drosophila. With enhanced sensitivity to subthreshold voltages, VARNAM resolves postsynaptic potentials in slices and cortical and hippocampal rhythms in freely behaving mice. Together, VARNAM lends a new hue to the optical toolbox, opening the door to high-speed in vivo multispectral functional imaging.

An Automated Microfluidic Assay for Photonic Crystal Enhanced Detection and Analysis of an Antiviral Antibody Cancer Biomarker in Serum.

We report on the implementation of an automated platform for detecting the presence of an antibody biomarker for human papillomavirus-associated oropharyngeal cancer from a single droplet of serum, in which a nanostructured photonic crystal surface is used to amplify the output of a fluorescence-linked immunosorbent assay. The platform is comprised of a microfluidic cartridge with integrated photonic crystal chips that interfaces with an assay instrument that automates the introduction of reagents, wash steps, and surface drying. Upon assay completion, the cartridge interfaces with a custom laser-scanning instrument that couples light into the photonic crystal at the optimal resonance condition for fluorescence enhancement. The instrument is used to measure the fluorescence intensity values of microarray spots corresponding to the biomarkers of interest, in addition to several experimental controls that verify correct functioning of the assay protocol. In this work, we report both dose-response characterization of the system using anti-E7 antibody introduced at known concentrations into serum and characterization of a set of clinical samples from which results were compared with a conventional enzyme-linked immunosorbent assay (ELISA) performed in microplate format. The demonstrated capability represents a simple, rapid, automated, and high-sensitivity method for multiplexed detection of protein biomarkers from a low-volume test sample.

Isolation, Detection, and Quantification of Cancer Biomarkers in HPV-Associated Malignancies.

Human Papillomavirus (HPV) infection has been recognized as the main etiologic factor in the development of various cancers including penile, vulva, oropharyngeal and cervical cancers. In the development of cancer, persistent HPV infections induce E6 and E7 oncoproteins, which promote cell proliferation and carcinogenesis resulting elevated levels of host antibodies (e.g., anti-HPV16 E7 antibody). Currently, these cancers are clinically diagnosed using invasive biopsy-based tests, which are performed only in centralized labs by experienced clinical staff using time-consuming and expensive tools and technologies. Therefore, these obstacles constrain their utilization at primary care clinics and in remote settings, where resources are limited. Here, we present a rapid, inexpensive, reliable, easy-to-use, customized immunoassay platform following a microfluidic filter device to detect and quantify anti-HPV16 E7 antibodies from whole blood as a non-invasive assisting technology for diagnosis of HPV-associated malignancies, especially, at primary healthcare and remote settings. The platform can detect and quantify anti-HPV16 E7 antibody down to 2.87 ng/mL. We further validated our immunoassay in clinical patient samples and it provided significantly high responses as compared to control samples. Thus, it can be potentially implemented as a pretesting tool to identify high-risk groups for broad monitoring of HPV-associated cancers in resource-constrained settings.

Monitoring Neutropenia for Cancer Patients at the Point of Care.

Neutrophils have a critical role in regulating the immune system. The immune system is compromised during chemotherapy, increasing infection risks and imposing a need for regular monitoring of neutrophil counts. Although commercial hematology analyzers are currently used in clinical practice for neutrophil counts, they are only available in clinics and hospitals, use large blood volumes, and are not available at the point of care (POC). Additionally, phlebotomy and blood processing require trained personnel, where patients are often admitted to hospitals when the infections are at late stage due to lack of frequent monitoring. Here, a reliable method is presented that selectively captures and quantifies white blood cells (WBCs) and neutrophils from a finger prick volume of whole blood by integrating microfluidics with high-resolution imaging algorithms. The platform is compact, portable, and easy to use. It captures and quantifies WBCs and neutrophils with high efficiency (>95%) and specificity (>95%) with an overall 4.2% bias compared to standard testing. The results from a small cohort of patients (N = 11 healthy, N = 5 lung and kidney cancer) present a unique disposable cell counter, demonstrating the ability of this tool to monitor neutrophil and WBC counts within clinical or in resource-constrained environments.

Photonic crystals: emerging biosensors and their promise for point-of-care applications.

Biosensors are extensively employed for diagnosing a broad array of diseases and disorders in clinical settings worldwide. The implementation of biosensors at the point-of-care (POC), such as at primary clinics or the bedside, faces impediments because they may require highly trained personnel, have long assay times, large sizes, and high instrumental cost. Thus, there exists a need to develop inexpensive, reliable, user-friendly, and compact biosensing systems at the POC. Biosensors incorporated with photonic crystal (PC) structures hold promise to address many of the aforementioned challenges facing the development of new POC diagnostics. Currently, PC-based biosensors have been employed for detecting a variety of biotargets, such as cells, pathogens, proteins, antibodies, and nucleic acids, with high efficiency and selectivity. In this review, we provide a broad overview of PCs by explaining their structures, fabrication techniques, and sensing principles. Furthermore, we discuss recent applications of PC-based biosensors incorporated with emerging technologies, including telemedicine, flexible and wearable sensing, smart materials and metamaterials. Finally, we discuss current challenges associated with existing biosensors, and provide an outlook for PC-based biosensors and their promise at the POC.

Advances in biosensing strategies for HIV-1 detection, diagnosis, and therapeutic monitoring.

HIV-1 is a major global epidemic that requires sophisticated clinical management. There have been remarkable efforts to develop new strategies for detecting and treating HIV-1, as it has been challenging to translate them into resource-limited settings. Significant research efforts have been recently devoted to developing point-of-care (POC) diagnostics that can monitor HIV-1 viral load with high sensitivity by leveraging micro- and nano-scale technologies. These POC devices can be applied to monitoring of antiretroviral therapy, during mother-to-child transmission, and identification of latent HIV-1 reservoirs. In this review, we discuss current challenges in HIV-1 diagnosis and therapy in resource-limited settings and present emerging technologies that aim to address these challenges using innovative solutions.