Re-Evaluating the Conventional Wisdom about Binding Assays.

Analytical technologies based on binding assays have evolved substantially since their inception nearly 60 years ago, but our conceptual understanding of molecular recognition has not kept pace. Contemporary technologies, such as single-molecule and digital measurements, have challenged, or even rendered obsolete, core concepts behind conventional binding assay design. Here, we explore the fundamental principles underlying molecular recognition systems, which we consider in terms of signals generated through concentration-dependent shifts in equilibrium. We challenge certain orthodoxies related to binding-based detection assays, including the primary importance of a low dissociation constant (KD) and the extent to which this parameter constrains dynamic range and limit of detection. Lastly, we identify key principles for designing binding assays that are optimally suited for a given detection application.

Comparing Sensor Fusion and Multimodal Chemometric Models for Monitoring U(VI) in Complex Environments Representative of Irradiated Nuclear Fuel.

Optical sensors and chemometric models were leveraged for the quantification of uranium(VI) (0-100 µg mL-1), europium (0-150 µg mL-1), samarium (0-250 µg mL-1), praseodymium (0-350 µg mL-1), neodymium (0-1000 µg mL-1), and HNO3 (2-4 M) with varying corrosion product (iron, nickel, and chromium) levels using laser fluorescence, Raman scattering, and ultraviolet-visible-near-infrared absorption spectra. In this paper, an efficient approach to developing and evaluating tens of thousands of partial least-squares regression (PLSR) models, built from fused optical spectra or multimodal acquisitions, is discussed. Each PLSR model was optimized with unique preprocessing combinations, and features were selected using genetic algorithm filters. The 7-factor D-optimal design training set contained just 55 samples to minimize the number of samples. The performance of PLSR models was evaluated by using an automated latent variable selection script. PLS1 regression models tailored to each species outperformed a global PLS2 model. PLS1 models built using fused spectra data and a multimodal (i.e., analyzed separately) approach yielded similar information, resulting in percent root-mean-square error of prediction values of 0.9-5.7% for the seven factors. The optical techniques and data processing strategies established in this study allow for the direct analysis of numerous species without measuring luminescence lifetimes or relying on a standard addition approach, making it optimal for near-real-time, in situ measurements. Nuclear reactor modeling helped bound training set conditions and identified elemental ratios of lanthanide fission products to characterize the burnup of irradiated nuclear fuel. Leveraging fluorescence, spectrophotometry, experimental design, and chemometrics can enable the remote quantification and characterization of complex systems with numerous species, monitor system performance, help identify the source of materials, and enable rapid high-throughput experiments in a variety of industrial processes and fundamental studies.

Effects of a nuclear-disturbed environment on electromagnetic wave propagation through the atmosphere.

This paper investigates the effects of a nuclear-disturbed environment on the transmission of electromagnetic (EM) waves through the atmosphere. An atmospheric nuclear detonation can produce heightened free electron densities in the surrounding atmosphere that can disrupt EM waves that propagate through the disturbed region. Radiation transport models simulated the ionization and free electron densities created in the atmosphere from a 1 MT detonation at heights of burst of 5 km, 25 km, and 75 km. Recombination rates for the free electrons in the atmosphere were applied, from previous work in the literature, to determine the nuclear-induced electron densities as a function of time and space after the detonation. A ray-tracing algorithm was applied to determine the refraction and reflection of waves propagating in the different nuclear-disturbed environments. The simulation results show that the free electron plasma created from an atmospheric nuclear detonation depend on the height of burst of the weapon, the weapon yield, and the time after detonation. Detonations at higher altitudes produce higher free electron densities for greater durations and over larger ranges. The larger the free electron densities, the greater the impact on EM wavelengths in regards to refraction, reflection, and absorption in the atmosphere. An analysis of modern infrastructure and the effects of nuclear-disturbed atmospheres on different signal wavelengths and systems is discussed.

Space-based quantum networking in the presence of a nuclear disturbed environment.

Space-based quantum networks provide a means for near-term long-distance transmission of quantum information. This article analyzed the performance of a downlink quantum network between a low-Earth-orbit satellite and an observatory operating in less-than-ideal atmospheric conditions. The effects from fog, haze, and a nuclear disturbed environment on the long-range distribution of quantum states were investigated. A density matrix that estimates the quantum state by capturing the effects from increased signal loss and elevated background noise to estimate the state fidelity of the transmitted quantum state was developed. It was found that the nuclear disturbed environment and other atmospheric effects have a degrading effect on the quantum state. These environments impede the ability to perform quantum communications for the duration of the effects. In the case of the nuclear disturbed environment, the nuclear effects subside quickly, and network performance should return to normal by the next satellite pass.

Effects of a nuclear disturbed environment on a quantum free space optical link.

This manuscript investigates the potential effect of a nuclear-disturbed atmospheric environment on the signal attenuation of a ground/satellite transmitter/receiver system for both classical optical and quantum communications applications. Attenuation of a signal transmitted through the rising nuclear cloud and the subsequently transported debris is modeled climatologically for surface-level detonations of 10 kt, 100 kt, and 1 Mt. Attenuation statistics were collected as a function of time after detonation. These loss terms were compared to normal loss sources such as clouds, smoke from fires, and clear sky operation. Finally, the loss was related to the degradation of transmitted entanglement derived from Bayesian mean estimation.

High-Fidelity Nanopore Sequencing of Ultra-Short DNA Targets.

Nanopore sequencing offers a portable and affordable alternative to sequencing-by-synthesis methods but suffers from lower accuracy and cannot sequence ultrashort DNA. This puts applications such as molecular diagnostics based on the analysis of cell-free DNA or single-nucleotide variants (SNVs) out of reach. To overcome these limitations, we report a nanopore-based sequencing strategy in which short target sequences are first circularized and then amplified via rolling-circle amplification to produce long stretches of concatemeric repeats. After sequencing on the Oxford Nanopore Technologies MinION platform, the resulting repeat sequences can be aligned to produce a highly accurate consensus that reduces the high error-rate present in the individual repeats. Using this approach, we demonstrate for the first time the ability to obtain unbiased and accurate nanopore data for target DNA sequences <100 bp. Critically, this approach is sensitive enough to achieve SNV discrimination in mixtures of sequences and even enables quantitative detection of specific variants present at ratios of <10%. Our method is simple, cost-effective, and only requires well-established processes. It therefore expands the utility of nanopore sequencing for molecular diagnostics and other applications, especially in resource-limited settings.

Thermodynamics of engineered gold binding peptides: establishing the structure-activity relationships.

Adsorption behavior of a gold binding peptide was experimentally studied to achieve kinetics and thermodynamics parameters toward understanding of the binding of an engineered peptide onto a solid metal surface. The gold-binding peptide, GBP1, was originally selected using a cell surface display library and contains 14 amino acid residues. In this work, single- and three-repeats of GBP1 were used to assess the effects of two parameters: molecular architecture versus secondary structure on adsorption on to gold substrate. The adsorption measurements were carried out using surface plasmon resonance (SPR) spectroscopy at temperatures ranging from 10 to 55 °C. At all temperatures, two different regimes of peptide adsorption were observed, which, based on the model, correspond to two sets of thermodynamics values. The values of enthalpy, ΔH(ads), and entropy, ΔS(ads), in these two regimes were determined using the van't Hoff approach and Gibbs-Helmholtz relationship. In general, the values of enthalpy for both peptides are negative indicating GBP1 binding to gold is an exothermic phenomenon and that the binding of three repeat gold binding peptide (3l-GBP1) is almost 5 times tighter than that for the single repeat (l-GBP1). More intriguing result is that the entropy of adsorption for the 3l-GBP1 is negative (-43.4 ± 8.5 cal/(mol K)), while that for the l-GBP1 is positive (10.90 ± 1.3 cal/(mol K)). Among a number of factors that synergistically contribute to the decrease of entropy, long-range ordered self-assembly of the 3l-GBP1 on gold surface is the most effective, probably through both peptide-solid and peptide-peptide intermolecular interactions. Additional adsorption experiments were conducted in the presence of 2,2,2-trifluoroethanol (TFE) to determine how the conformational structures of the biomolecules responded to the environmental perturbation. We found that the peptides differ in their conformational responses to the change in solution conditions; while l-GBP does not fold in the presence of TFE, 3l-GBP1 adopted two types of secondary structure (ß-strand, α-helix) and that peptide's binding to the solid is enhanced by the presence of low percentages of TFE solvent. Not only do these kinetics and thermodynamics results provide adsorption behavior and binding of genetically engineered peptides for inorganics (GEPI), but they could also provide considerable insights into fundamental understanding peptide molecular recognition and their selective specificity for the solids. Moreover, comprehensive work described herein suggests that multiple repeat forms of the solid binding peptides possess a conformational component that can be exploited to further tailor affinity and binding of a given sequence to a solid material followed by ordered assembly as a convenient tool in future practical applications.

Chest X-Ray Remains a Vital Component Prior to Tube Thoracostomy.

INTRODUCTION: The identification and treatment of traumatic pneumothorax (PTX) has long been a focus of bedside imaging in the trauma patient. While the emergence of bedside ultrasound (BUS) provides an opportunity for earlier detection, the need for tube thoracostomy (TT) based on bedside imaging, including BUS and supine AP chest X-ray (CXR) is less established in the medical literature. METHODS: Retrospective data from 2017 to 2020 were collected of all adult trauma activations at a level 1 rural trauma facility. Every adult patient included in this study received a CXR and BUS (eFast) upon arrival. The need for TT was determined by the emergency medicine attending or the trauma surgery attending evaluating the patient. McNemar's chi-squared test and conditional logistic regression analysis were performed comparing BUS, CXR, and the combination of BUS and CXR findings for the need for TT. Subgroup analyses were performed comparing BUS, CXR, and the combination of BUS and CXR for the detection of PTX compared to CT scan. RESULTS: Of the 12,244 patients who underwent trauma activation during this timeframe, 602 were included in the study. 74.9% were males with an age range of 36-63 years. Of the 602 patients, 210 received TT. Positive PTX was recorded with BUS in 128 (21%) patients with 16 false negatives (FNs) and 98 false positives (FPs), 100 (17%) PTX were identified with CXR with 114 FNs and 4 FPs, and 72 (11.9%) were noted on both CXR and BUS with 140 FNs and 2 FPs. The odds ratio of TT placement was 22 times with positive BUS alone (P < .0001, 95% CI: 10.9-43.47), 47 times with positive CXR alone (P < .0001, 95% CI: 16.99-127.5), and 70 times with both positive CXR and BUS (P < .0001, 95% CI: 17.08-288.4). CONCLUSION: A positive finding of PTX on BUS combined with CXR is more indicative of the need for TT in the trauma patient when compared with BUS or CXR alone.

Neuropsychiatric Complications of Hypervitaminosis D: Diagnosis, Evaluation, and Treatment.

The Psychiatric Consultation Service at Massachusetts General Hospital sees medical and surgical inpatients with comorbid psychiatric symptoms and conditions. During their twice-weekly rounds, Dr Stern and other members of the Consultation Service discuss diagnosis and management of hospitalized patients with complex medical or surgical problems who also demonstrate psychiatric symptoms or conditions. These discussions have given rise to rounds reports that will prove useful for clinicians practicing at the interface of medicine and psychiatry.Prim Care Companion CNS Disord 2024;26(3):23f03680. Author affiliations are listed at the end of this article.

Surgical Management of Extravaginal Testicular Torsion in a 16-Year-Old Male: A Case Report.

Extravaginal torsion (EVT) is a rare type of testicular torsion that usually occurs in neonates. The primary type of testicular torsion that occurs in adolescents is intravaginal torsion. In this case report, we describe the first case of EVT reported in a 16-year-old male with a contralateral bell clapper deformity and subsequent surgical management using a tunica vaginalis flap and bilateral orchiopexy. In discussion of this case, we examine possible anatomical causes of EVT and suggestions for appropriate surgical management.

Spaceflight effects on human vascular smooth muscle cell phenotype and function.

The cardiovascular system is strongly impacted by the hazards of spaceflight. Astronauts spending steadily increasing lengths of time in microgravity are subject to cardiovascular deconditioning resulting in loss of vascular tone, reduced total blood volume, and diminished cardiac output. Appreciating the mechanisms by which the cells of the vasculature are altered during spaceflight will be integral to understanding and combating these deleterious effects as the human presence in space advances. In this study, we performed RNA-Seq analysis coupled with review by QIAGEN Ingenuity Pathway Analysis software on human aortic smooth muscle cells (HASMCs) cultured for 3 days in microgravity and aboard the International Space Station to assess the transcriptomic changes that occur during spaceflight. The results of our RNA-Seq analysis show that SMCs undergo a wide range of transcriptional alteration while in space, significantly affecting 4422 genes. SMCs largely down-regulate markers of the contractile, synthetic, and osteogenic phenotypes including smooth muscle alpha actin (αSMA), matrix metalloproteinases (MMPs), and bone morphogenic proteins (BMPs). Additionally, components of several cellular signaling pathways were strongly impacted including the STAT3, NFκB, PI3K/AKT, HIF1α, and Endothelin pathways. This study highlights the significant changes in transcriptional behavior SMCs exhibit during spaceflight and puts these changes in context to better understand vascular function in space.

Non-cutaneous presentation of mycosis fungoides involving the penile corpora: Case report and review of current literature.

Mycosis fungoides involvement of genitalia is rare. We present a 63-year-old man with history of cutaneous T cell lymphoma with large cell transformation status post multiple electron beam radiation cycles who presented with a new, enlarging penile mass. He underwent ultrasound, MRI, and excisional biopsy. Pathological results indicated hematogenous spread of T cell lymphoma with large cell transformation. Peri-operative radiation was performed, and the patient had significant reduction in penile mass size but some subsequent erectile dysfunction. In discussion of this case, we examine management of penile mycosis fungoides.

Myoepithelial carcinoma of the bladder: Case report and review of the literature.

Myoepithelial carcinoma is a neoplasm that classically arises in the parotid glands, nasopharynx, paranasal sinus, and nasal cavity of the head and neck. It rarely arises in other organs or soft tissues and involvement of genitourinary organs is distinctly rare. We describe a case of a 21-year-old male, presenting with nausea, weight loss, and worsening suprapubic pain over 3 months, found to have a large mass at the dome of the bladder. Partial cystectomy was ultimately performed revealing myoepithelial carcinoma of the bladder. The patient is free of disease at four years without the need for systemic therapy.

Extending the dynamic range of biomarker quantification through molecular equalization.

Precision medicine requires highly scalable methods of multiplexed biomarker quantification that can accurately describe patient physiology. Unfortunately, contemporary molecular detection methods are generally limited to a dynamic range of sensitivity spanning just 3-4 orders of magnitude, whereas the actual physiological dynamic range of the human plasma proteome spans more than 10 orders of magnitude. Current methods rely on sample splitting and differential dilution to compensate for this mismatch, but such measures greatly limit the reproducibility and scalability that can be achieved-in particular, the effects of non-linear dilution can greatly confound the analysis of multiplexed assays. We describe here a two-pronged strategy for equalizing the signal generated by each analyte in a multiplexed panel, thereby enabling simultaneous quantification of targets spanning a wide range of concentrations. We apply our 'EVROS' strategy to a proximity ligation assay and demonstrate simultaneous quantification of four analytes present at concentrations spanning from low femtomolar to mid-nanomolar levels. In this initial demonstration, we achieve a dynamic range spanning seven orders of magnitude in a single 5 µl sample of undiluted human serum, highlighting the opportunity to achieve sensitive, accurate detection of diverse analytes in a highly multiplexed fashion.

Fabrication of hierarchical hybrid structures using bio-enabled layer-by-layer self-assembly.

Development of versatile and flexible assembly systems for fabrication of functional hybrid nanomaterials with well-defined hierarchical and spatial organization is of a significant importance in practical nanobiotechnology applications. Here we demonstrate a bio-enabled self-assembly technique for fabrication of multi-layered protein and nanometallic assemblies utilizing a modular gold-binding (AuBP1) fusion tag. To accomplish the bottom-up assembly we first genetically fused the AuBP1 peptide sequence to the C'-terminus of maltose-binding protein (MBP) using two different linkers to produce MBP-AuBP1 hetero-functional constructs. Using various spectroscopic techniques, surface plasmon resonance (SPR) and localized surface plasmon resonance (LSPR), we verified the exceptional binding and self-assembly characteristics of AuBP1 peptide. The AuBP1 peptide tag can direct the organization of recombinant MBP protein on various gold surfaces through an efficient control of the organic-inorganic interface at the molecular level. Furthermore using a combination of soft-lithography, self-assembly techniques and advanced AuBP1 peptide tag technology, we produced spatially and hierarchically controlled protein multi-layered assemblies on gold nanoparticle arrays with high molecular packing density and pattering efficiency in simple, reproducible steps. This model system offers layer-by-layer assembly capability based on specific AuBP1 peptide tag and constitutes novel biological routes for biofabrication of various protein arrays, plasmon-active nanometallic assemblies and devices with controlled organization, packing density and architecture.

Lessons on Increasing Racial and Health Equity From Accountable Health Communities.

BACKGROUND: The Accountable Health Communities model (AHC) was developed to test whether systematically screening for health-related social needs and referrals to community-based organizations to resolve unmet needs would affect healthcare use and costs for CMS beneficiaries. Purpose: The AHC model required applicants to develop Disparities Impact Statements (DIS), to increase the model's potential impact on health equity. METHODS: Authors conducted a thematic analysis of awardees' DISs to identify minority and underserved populations of focus, and the strategies awardees used to increase equitable participation in the model by minority and underserved populations. RESULTS: Most awardees focused on multiple minority and underserved populations and used multipronged innovative strategies to pursue equity goals. CONCLUSIONS: Considering recent health equity advancements as Executive Order 13985 and the release of CMS Innovation Center's Strategy Refresh, with highlights of health equity best practices from the AHC model, assessing use of DISs in the AHC model provides valuable lessons. Implications: Given HHS' broadscale promotion of DISs adoption as a viable quality improvement approach to achieving health equity, disseminating how the tool was used by a myriad of organizational types in the AHC model is critically important to improving future efforts to increase equity.

Pre-equilibrium biosensors as an approach towards rapid and continuous molecular measurements.

Almost all biosensors that use ligand-receptor binding operate under equilibrium conditions. However, at low ligand concentrations, the equilibration with the receptor (e.g., antibodies and aptamers) becomes slow and thus equilibrium-based biosensors are inherently limited in making measurements that are both rapid and sensitive. In this work, we provide a theoretical foundation for a method through which biosensors can quantitatively measure ligand concentration before reaching equilibrium. Rather than only measuring receptor binding at a single time-point, the pre-equilibrium approach leverages the receptor's kinetic response to instantaneously quantify the changing ligand concentration. Importantly, by analyzing the biosensor output in frequency domain, rather than in the time domain, we show the degree to which noise in the biosensor affects the accuracy of the pre-equilibrium approach. Through this analysis, we provide the conditions under which the signal-to-noise ratio of the biosensor can be maximized for a given target concentration range and rate of change. As a model, we apply our theoretical analysis to continuous insulin measurement and show that with a properly selected antibody, the pre-equilibrium approach could make the continuous tracking of physiological insulin fluctuations possible.

Accelerated Electron Transfer in Nanostructured Electrodes Improves the Sensitivity of Electrochemical Biosensors.

Electrochemical biosensors hold the exciting potential to integrate molecular detection with signal processing and wireless communication in a miniaturized, low-cost system. However, as electrochemical biosensors are miniaturized to the micrometer scale, their signal-to-noise ratio degrades and reduces their utility for molecular diagnostics. Studies have reported that nanostructured electrodes can improve electrochemical biosensor signals, but since the underlying mechanism remains poorly understood, it remains difficult to fully exploit this phenomenon to improve biosensor performance. In this work, electrochemical aptamer biosensors on nanoporous electrode are optimized to achieve improved sensitivity by tuning pore size, probe density, and electrochemical measurement parameters. Further, a novel mechanism in which electron transfer is physically accelerated within nanostructured electrodes due to reduced charge screening, resulting in enhanced sensitivity is proposed and experimentally validated. In concert with the increased surface areas achieved with this platform, this newly identified effect can yield an up to 24-fold increase in signal level and nearly fourfold lower limit of detection relative to planar electrodes with the same footprint. Importantly, this strategy can be generalized to virtually any electrochemical aptamer sensor, enabling sensitive detection in applications where miniaturization is a necessity, and should likewise prove broadly applicable for improving electrochemical biosensor performance in general.

Upgrading on radical prostatectomy specimens of very low- and low-risk prostate cancer patients on active surveillance: A population-level analysis.

INTRODUCTION: A proportion of prostate cancer (PCa) patients initially managed with active surveillance (AS) are upgraded to a higher Gleason score (GS) at the time of radical prostatectomy (RP). Our objective was to determine predictors of upgrading on RP specimens using a national database. METHODS: The Surveillance, Epidemiology, and End Results Prostate with Watchful Waiting database was used to identify AS patients diagnosed with very low- or low-risk PCa who underwent delayed RP between 2010 and 2015. The primary outcome was upgrading to GS 7 disease or worse. Logistic regression analyses were used to evaluate demographic and oncological predictors of upgrading on final specimen. RESULTS: A total of 3775 men underwent RP after a period of AS, 3541 (93.8%) of whom were cT2a; 792 (21.0%) patients were upgraded on RP specimen, with 85.4%, 10.6%, and 3.4% upgraded to GS 7(3+4), 7(4+3), and 8 diseases, respectively. On multivariable analysis, higher prostate-specific antigen (PSA) at diagnosis (5-10 vs. 0-2 ng/ml, odd ratio [OR] 2.59, p<0.001) and percent core involvement (80-100% vs. 0-20%, OR 2.52, p=0.003) were significant predictors of upgrading on final RP specimen, whereas higher socioeconomic status predicted lower odds of upgrading (highest vs. lowest quartile OR 0.75, p=0.013). CONCLUSIONS: Higher baseline PSA and percent positive cores involvement are associated with significantly increased risk of upgrading on RP after AS, whereas higher socioeconomic status predicts lower odds of such events. These results may help identify patients at increased risk of adverse pathology on final specimen who may benefit from earlier definitive treatment.