Two simplified tooth sample preparation methods for conventional laboratory and RapidHIT™ ID workflows.

Disaster victim identification (DVI) refers to the forensic identification of unknown individuals following a mass disaster event. Human dental structures can contain viable DNA sources when other soft tissues are compromised. However, labor-intensive sample preparation performed by extensively trained personnel is needed to expose the nuclear material for traditional forensic DNA workflows.

Comprehensive performance evaluation of ligand-binding assay-LC-MS/MS method for co-dosed monoclonal anti-SARS-CoV-2 antibodies (AZD7442).

AZD7442 is a combination SARS-CoV-2 therapy comprising two co-dosed monoclonal antibodies. The authors validated a hybrid ligand-binding assay-LC-MS/MS method for pharmacokinetic assessment of AZD7442 in human serum with nominal concentration range of each analyte of 0.300-30.

Acidic Methanol Treatment Facilitates Matrix-Assisted Laser Desorption Ionization-Mass Spectrometry Imaging of Energy Metabolism.

Detection of small molecule metabolites (SMM), particularly those involved in energy metabolism using MALDI-mass spectrometry imaging (MSI), is challenging due to factors including ion suppression from other analytes present (e.g., proteins and lipids).

Digital image analysis for biothreat detection rapid centrifugal microfluidic orthogonal flow immunocapture.

We report clear proof-of-principle for centrifugally-driven, multiplexed, paper-based orthogonal flow sandwich-style immunocapture (cOFI) and colorimetric detection of Zaire Ebola virus-like particles. Capture antibodies are immobilized onto nanoporous nitrocellulose membranes that are then laminated into polymeric microfluidic discs to yield ready-to-use analytical devices. Fluid flow is controlled solely by rotational speed, obviating the need for complex pneumatic pumping systems, and providing more precise flow control than with the capillary-driven flow used in traditional lateral flow immunoassays (LFIs).

Towards an affinity-free, centrifugal microfluidic system for rapid, automated forensic differential extraction.

Biological evidence originating from victims of sexual assault is often comprised of unbalanced cellular mixtures with significantly higher contributions from the victim's genetic material. Enrichment of the forensically-critical sperm fraction (SF) with single-source male DNA relies on differential extraction (DE), a manually-intensive process that is prone to contamination. Due to DNA losses from sequential washing steps, some existing DE methods often fail to generate sufficient sperm cell DNA recovery for perpetrator(s) identification.

Cell-type specific profiling of histone post-translational modifications in the adult mouse striatum.

Epigenetic gene regulation in the heterogeneous brain remains challenging to decipher with current strategies. Bulk tissue analysis from pooled subjects reflects the average of cell-type specific changes across cell-types and individuals, which obscures causal relationships between epigenetic modifications, regulation of gene expression, and complex pathology. To address these limitations, we optimized a hybrid protocol, ICuRuS, for the isolation of nuclei tagged in specific cell-types and histone post translational modification profiling from the striatum of a single mouse.

Cocaine regulation of Nr4a1 chromatin bivalency and mRNA in male and female mice.

Cocaine epigenetically regulates gene expression via changes in histone post-translational modifications (HPTMs). We previously found that the immediate early gene Nr4a1 is epigenetically activated by cocaine in mouse brain reward regions. However, few studies have examined multiple HPTMs at a single gene.

A novel method for inward fluid displacement in centrifugal microdevices for highly integrated nucleic acid processing with long-term reagent storage.

Rotationally-driven lab-on-a-disc (LoaD) microfluidic systems are among the most promising methods for realizing complex nucleic acid (NA) testing at the point-of-need (PoN). However, despite significant advancements in NA amplification methods, very few sample-to-answer centrifugal microfluidic platforms have been realized due, in part, to a lack of on-disc sample preparation. In many instances, NA extraction (NAE) and/or lysis must be performed off-disc using conventional laboratory equipment and methods, thus tethering the assay to centralized facilities.

Characterization of a Centrifugal Microfluidic Orthogonal Flow Platform.

To bring to bear the power of centrifugal microfluidics on vertical flow immunoassays, control of flow orthogonally through nanoporous membranes is essential. The on-disc approach described here leverages the rapid print-cut-laminate (PCL) disc fabrication and prototyping method to create a permanent seal between disc materials and embedded nanoporous membranes. Rotational forces drive fluid flow, replacing capillary action, and complex pneumatic pumping systems.

A Membrane-Modulated Centrifugal Microdevice for Enzyme-Linked Immunosorbent Assay-Based Detection of Illicit and Misused Drugs.

Increased opioid use and misuse have imposed large analytical demands across clinical and forensic sectors. Due to the absence of affordable, accurate, and simple on-site tests (e.g.

Multiplexed Centrifugal Microfluidic System for Dynamic Solid-Phase Purification of Polynucleic Acids Direct from Buccal Swabs.

This report describes the development of a centrifugally controlled microfluidic dynamic solid-phase extraction (dSPE) platform to reliably obtain amplification-ready nucleic acids (NAs) directly from buccal swab cuttings. To our knowledge, this work represents the first centrifugal microdevice for comprehensive preparation of high-purity NAs from raw buccal swab samples. Direct-from-swab cellular lysis was integrated upstream of NA extraction, and automatable laser-controlled on-board microvalving strategies provided the strict spatiotemporal fluidic control required for practical point-of-need use.

Digital postprocessing and image segmentation for objective analysis of colorimetric reactions.

Recently, there has been an explosion of scientific literature describing the use of colorimetry for monitoring the progression or the endpoint result of colorimetric reactions. The availability of inexpensive imaging technology (e.g.

Closable Valves and Channels for Polymeric Microfluidic Devices.

This study explores three unique approaches for closing valves and channels within microfluidic systems, specifically multilayer, centrifugally driven polymeric devices. Precise control over the cessation of liquid movement is achieved through either the introduction of expanding polyurethane foam, the application of direct contact heating, or the redeposition of xerographic toner via chloroform solvation and evaporation. Each of these techniques modifies the substrate of the microdevice in a different way.

Optically-controlled closable microvalves for polymeric centrifugal microfluidic devices.

Microvalving is a pivotal component in many microfluidic lab-on-a-chip platforms and micro-total analysis systems (μTAS). Effective valving is essential for the integration of multiple unit operations, such as, liquid transport, mixing, aliquoting, metering, washing, and fractionation. The ideal microfluidic system integrates numerous, sequential unit operations, provides precise spaciotemporal reagent release and flow control, and is amenable to rapid, low-cost fabrication and prototyping.

A survey of bacteria associated with various life stages of primary colonizers: Lucilia sericata and Phormia regina.

Blow flies are common primary colonizers of carrion, play an important role in the transfer of microbes between environments, and serve as a vector for many human pathogens. While some investigation has begun regarding the bacteria associated with different life stages of blow flies, a well replicated study is currently not available for the majority of blow flies. This study investigated bacteria associated with successive life stages of blow fly species Lucilia sericata and Phormia regina.

Real Time Electronic Feedback for Improved Acoustic Trapping of Micron-Scale Particles.

Acoustic differential extraction has been previously reported as a viable alternative to the repetitive manual pipetting and centrifugation steps for isolating sperm cells from female epithelial cells in sexual assault sample evidence. However, the efficiency of sperm cell isolation can be compromised in samples containing an extremely large number of epithelial cells. When highly concentrated samples are lysed, changes to the physicochemical nature of the medium surrounding the cells impacts the acoustic frequency needed for optimal trapping.

A eukaryotic community succession based method for postmortem interval (PMI) estimation of decomposing porcine remains.

Recent, short-term studies on porcine and human models (albeit with few replicates) demonstrated that the succession of the microbial community of remains may be used to estimate time since death. Using a porcine model (N=6) over an extended period of time (1703 ADD, or two months), this study characterized the eukaryote community of decomposing remains. Skin microbial samples were collected from the torso of each set of remains every day during the first week, on alternate days during the second week, and once a week for the remainder of the 60-day period; all collection intervals were recorded in accumulated degree days (ADD).

Evaluation of Eph receptor and ephrin expression within the human cornea and limbus.

Eph receptor tyrosine kinases and their ligands, the ephrins, regulate the development and maintenance of multiple organs but little is known about their potential role within the cornea. The purpose of this study was to perform a thorough investigation of Eph/ephrin expression within the human cornea including the limbal stem cell niche. Initially, immunohistochemistry was performed on human donor eyes to determine the spatial distribution of Eph receptors and ephrins in the cornea and limbus.

Genetic rescue and inbreeding depression in Mexican wolves.

Although inbreeding can reduce individual fitness and contribute to population extinction, gene flow between inbred but unrelated populations may overcome these effects. Among extant Mexican wolves (Canis lupus baileyi), inbreeding had reduced genetic diversity and potentially lowered fitness, and as a result, three unrelated captive wolf lineages were merged beginning in 1995. We examined the effect of inbreeding and the merging of the founding lineages on three fitness traits in the captive population and on litter size in the reintroduced population.

Disease-associated casein kinase I delta mutation may promote adenomatous polyps formation via a Wnt/beta-catenin independent mechanism.

The Wnt signaling pathway is critical for embryonic development and is dysregulated in multiple cancers. Two closely related isoforms of casein kinase I (CKIdelta and epsilon) are positive regulators of this pathway. We speculated that mutations in the autoinhibitory domain of CKIdelta/epsilon might upregulate CKIdelta/epsilon activity and hence Wnt signaling and increase the risk of adenomatous polyps and colon cancer.

An opposite role for tau in circadian rhythms revealed by mathematical modeling.

Biological clocks with a period of approximately 24 h (circadian) exist in most organisms and time a variety of functions, including sleep-wake cycles, hormone release, bioluminescence, and core body temperature fluctuations. Much of our understanding of the clock mechanism comes from the identification of specific mutations that affect circadian behavior. A widely studied mutation in casein kinase I (CKI), the CKIepsilon(tau) mutant, has been shown to cause a loss of kinase function in vitro, but it has been difficult to reconcile this loss of function with the current model of circadian clock function.

Control of mammalian circadian rhythm by CKIepsilon-regulated proteasome-mediated PER2 degradation.

The mammalian circadian regulatory proteins PER1 and PER2 undergo a daily cycle of accumulation followed by phosphorylation and degradation. Although phosphorylation-regulated proteolysis of these inhibitors is postulated to be essential for the function of the clock, inhibition of this process has not yet been shown to alter mammalian circadian rhythm. We have developed a cell-based model of PER2 degradation.