\u27Living cantilever arrays\u27 for characterization of mass of single live cells in fluids

The size of a cell is a fundamental physiological property and is closely regulated by various environmental and genetic factors. Optical or confocal microscopy can be used to measure the dimensions of adherent cells, and Coulter counter or flow cytometry ( forward scattering light intensity) can be used to estimate the volume of single cells in a flow. Although these methods could be used to obtain the mass of single live cells, no method suitable for directly measuring the mass of single adherent cells without detaching them from the surface is currently available. We report the design, fabrication, and testing of \u27living cantilever arrays\u27, an approach to measure the mass of single adherent live cells in fluid using silicon cantilever mass sensor. HeLa cells were injected into microfluidic channels with a linear array of functionalized silicon cantilevers and the cells were subsequently captured on the cantilevers with positive dielectrophoresis. The captured cells were then cultured on the cantilevers in a microfluidic environment and the resonant frequencies of the cantilevers were measured. The mass of a single HeLa cell was extracted from the resonance frequency shift of the cantilever and was found to be close to the mass value calculated from the cell density from the literature and the cell volume obtained from confocal microscopy. This approach can provide a new method for mass measurement of a single adherent cell in its physiological condition in a non-invasive manner, as well as optical observations of the same cell. We believe this technology would be very valuable for single cell time-course studies of adherent live cells

AN ESSENTIAL TOOL TO BE OPTIMIZED: SYNDROMIC MANAGEMENT OF VAGINAL DISCHARGE IN HAITI

Background: Haiti, like many low-income countries in crisis, has limited resources for etiologic diagnosis of vaginal discharge. As such, we sought to characterize variability in diagnoses of women presenting with vaginal discharge syndrome, with the goal to improve standardization of syndromic management. Materials and Methods: Participants aged 18 years and older endorsing vaginitis, or dysuria were recruited at Jerusalem Clinic over two, one-week periods in April 2018 and July 2019. We calculated Spearman rank correlations among history, exam findings, and diagnoses based on clinical presentation, to understand presentation groupings and their management. Results: Among 98 women, median age was 33.5 years, and most frequent symptoms were: vaginal discharge (97%), vaginal itch (73%), and/or suprapubic pain (68%). Most common physical exam findings were vaginal discharge (86%), suprapubic/lower quadrant tenderness (29%), cervical motion tenderness (24%), and cervical erythema (20%). Most symptoms and physical exam findings were weakly correlated with each other. Nearly one-third (31%) were diagnosed with normal physiologic vaginal discharge or no diagnosis, followed by Bacterial vaginosis (31%), vulvovaginal candidiasis (15%), cervicitis/PID (13%), and STI (7%). No reported symptoms strongly differentiated diagnostic categories. Diagnoses varied considerably by exam findings. Conclusions: The weak correlations between symptoms, exam findings, and diagnoses could represent variability in assessment. In the absence of reliable and accessible laboratory testing, the importance of standardizing syndromic management becomes increasingly relevant. Results from our study support the utility of speculum examination and more standardized documentation of physical exam findings. Next steps include the development of local algorithms to promote standardization of treatment of vaginal discharge syndrome

Portable Bacterial Identification System Based on Elastic Light Scatter Patterns

Background Conventional diagnosis and identification of bacteria requires shipment of samples to a laboratory for genetic and biochemical analysis. This process can take days and imposes significant delay to action in situations where timely intervention can save lives and reduce associated costs. To enable faster response to an outbreak, a low-cost, small-footprint, portable microbial-identification instrument using forward scatterometry has been developed. Results This device, weighing 9 lb and measuring 12 × 6 × 10.5 in., utilizes elastic light scatter (ELS) patterns to accurately capture bacterial colony characteristics and delivers the classification results via wireless access. The overall system consists of two CCD cameras, one rotational and one translational stage, and a 635-nm laser diode. Various software algorithms such as Hough transform, 2-D geometric moments, and the traveling salesman problem (TSP) have been implemented to provide colony count and circularity, centering process, and minimized travel time among colonies. Conclusions Experiments were conducted with four bacteria genera using pure and mixed plate and as proof of principle a field test was conducted in four different locations where the average classification rate ranged between 95 and 100%

Factors necessary to produce basoapical polarity in human glandular epithelium formed in conventional and high-throughput three-dimensional culture: example of the breast epithelium

<p>Abstract</p> <p>Background</p> <p>Basoapical polarity in epithelia is critical for proper tissue function, and control of proliferation and survival. Cell culture models that recapitulate epithelial tissue architecture are invaluable to unravel developmental and disease mechanisms. Although factors important for the establishment of basal polarity have been identified, requirements for the formation of apical polarity in three-dimensional tissue structures have not been thoroughly investigated.</p> <p>Results</p> <p>We demonstrate that the human mammary epithelial cell line-3522 S1, provides a resilient model for studying the formation of basoapical polarity in glandular structures. Testing three-dimensional culture systems that differ in composition and origin of substrata reveals that apical polarity is more sensitive to culture conditions than basal polarity. Using a new high-throughput culture method that produces basoapical polarity in glandular structures without a gel coat, we show that basal polarity-mediated signaling and collagen IV are both necessary for the development of apical polarity.</p> <p>Conclusion</p> <p>These results provide new insights into the role of the basement membrane, and especially collagen IV, in the development of the apical pole, a critical element of the architecture of glandular epithelia. Also, the high-throughput culture method developed in this study should open new avenues for high-content screening of agents that act on mammary tissue homeostasis and thus, on architectural changes involved in cancer development.</p

Mouse and human islets survive and function after coating by biosilicification

Inorganic materials have properties that can be advantageous in bioencapsulation for cell transplantation. Our aim was to engineer a hybrid inorganic/soft tissue construct by inducing pancreatic islets to grow an inorganic shell. We created pancreatic islets surrounded by porous silica, which has potential application in the immunoprotection of islets in transplantation therapies for type 1 diabetes. The new method takes advantage of the islet capsule surface as a template for silica formation. Mouse and human islets were exposed to medium containing saturating silicic acid levels for 9-15 min. The resulting tissue constructs were then cultured for up to 4 wk under normal conditions. Scanning electron microscopy and energy dispersive X-ray spectroscopy was used to monitor the morphology and elemental composition of the material at the islet surface. A cytokine assay was used to assess biocompatibility with macrophages. Islet survival and function were assessed by confocal microscopy, glucose-stimulated insulin release assays, oxygen flux at the islet surface, expression of key genes by RT-PCR, and syngeneic transplant into diabetic mice

Differential Mitochondrial Toxicity Screening and Multi- Parametric Data Analysis

Early evaluation of new drug entities for their potential to cause mitochondrial dysfunction is becoming an important task for drug development. Multi-parametric high-content screening (mp-HCS) of mitochondrial toxicity holds promise as a lead in-vitro strategy for drug testing and safety evaluations. In this study, we have developed a mp-HCS and multi-parametric data analysis scheme for assessing cell responses to induced mitochondrial perturbation. The mp-HCS measurements are shown to be robust enough to allow for quantitative comparison of biological systems with different metabolic pathways simulated by alteration of growth media. Substitution of medium glucose for galactose sensitized cells to drug action and revealed novel response parameters. Each compound was quantitatively characterized according to induced phenotypic changes of cell morphology and functionality measured by fluorescent biomarkers for mitochondrial activity, plasma membrane permeability, and nuclear morphology. Descriptors of drug effects were established by generation of a SCRIT (Specialized-Cell-Response-to-Induced-Toxicity) vector, consisting of normalized statistical measures of each parameter at each dose and growth condition. The dimensionality of SCRIT vectors depends on the number of parameters chosen, which in turn depends on the hypothesis being tested. Specifically, incorporation of three parameters of response into SCRIT vectors enabled clustering of 84 training compounds with known pharmacological and toxicological activities according to the degree of toxicity and mitochondrial involvement. Inclusion of 6 parameters enabled the resolution of more subtle differences between compounds within a common therapeutic class; scoring enabled a ranking of statins in direct agreement with clinical outcomes. Comparison of drug-induced changes required variations in glucose for separation of mitochondrial dysfunction from other types of cytotoxicity. These results also demonstrate that the number of drugs in a training set, the choice of parameters used in analysis, and statistical measures are fundamental for specific hypothesis testing and assessment of quantitative phenotypic differences

Comparative three-dimensional imaging of living neurons with confocal and atomic force microscopy

Atomic force microscopy applications extend across a number of fields; however, limitations have reduced its effectiveness in live cell analysis. This report discusses the use of AFM to evaluate the three-dimensional (3-D) architecture of living chick dorsal root ganglia and sympathetic ganglia. These data sets were compared to similar images acquired with confocal laser scanning microscopy of identical cells. For this comparison we made use of visualization techniques which were applicable to both sets of data and identified several issues when coupling these technologies. These direct comparisons offer quantitative validation and confirmation of the character of novel images acquired by AFM. This paper is one in a series emphasizing various new applications of AFM in neurobiology

Improving Domestic Tourism In Namibia

This report, commissioned by the Namibia Tourism Board, investigates domestic tourism in Namibia. Through analysis of archival research, observations, questionnaires, and interviews conducted with domestic tourists and service providers, findings on current travel trends, perceptions of tourism, and areas for improvement were developed. A set of recommendations was compiled for the Namibia Tourism Board to identify areas for further research and ultimately inform improvements to develop domestic tourism in Namibia to a successful and sustainable level

Does Education Influence Ethical Decisions? An International Study

This study examined whether having attended a public, private or religious affiliated grade and/or high school influenced a college student\u27s ethical decision making process. We also examined whether having taken an ethics course in college influences a student\u27s ethical decision making process. Our sample included 508 accounting students (237 men and 271 women) from Albania, Ecuador, Ireland and the United States. Our analyses indicated no differences in ethical decision making that associated with either grade-or-high-school education. While our data showed no difference in the reported attitudes between students from Ecuador and the United States after controlling for social desirability response bias, we found significant differences between the attitudes students from the United States and students from both Albania and Ireland. While gender was also significant for six of our seven scenarios, social desirability response bias was significant in all of our scenarios. © 2011 Springer Science+Business Media B.V