The laser print, cut, and laminate (PCL) method for microfluidic device fabrication can be leveraged for rapid and inexpensive prototyping of electrophoretic microchips useful for optimizing ...separation conditions. The rapid prototyping capability allows the evaluation of fluidic architecture, applied fields, reagent concentrations, and sieving matrix, all within the context of using fluorescence‐compatible substrates. Cyclic olefin copolymer and toner‐coated polyethylene terephthalate (tPeT) were utilized with the PCL technique and bonding methods optimized to improve device durability during electrophoresis. A series of separation channel designs and centrifugation conditions that provided successful loading of sieving polymer in less than 3 min was described. Separation of a 400‐base DNA sizing ladder provided calculated base resolution between 3 and 4 bases, a greater than 18‐fold improvement over separations on similar substrates. Finally, the accuracy and precision capabilities of these devices were demonstrated by separating and sizing DNA fragments of 147 and 167 bases as 148.62 ± 2 and 166.48 ± 3 bases, respectively.
Hematocrit (HCT) measurements are important clinical diagnostic variables that help physicians diagnose and treat various medical conditions, ailments, and diseases. In this work, we present the HCT ...Disc, a centrifugal microdevice fabricated by a Print, Cut and Laminate (PCL) method to generate a 12-sample HCT device from materials costing <0.5 USD (polyester and toner or PeT). Following introduction from a drop of blood (finger stick), whole blood metering and cell sedimentation are controlled by centrifugal force, only requiring a CD player motor as external hardware and, ultimately, a cell phone for detection. The sedimented volume from patient blood in the HCT Disc was analyzed using a conventional scanner/custom algorithm for analysis of the image to determine a hematocrit value, and these were compared to values generated in a clinical laboratory, which correlated well. To enhance portability and assure simplicity of the HCT measurement, values from image analysis by a cell phone using a custom application was compared to the scanner. Fifteen samples were analyzed with cell phone image analysis system and were found to be within 4% of the HCT values determined in the clinical lab. We demonstrate the feasibility of the PeT device for HCT measurement, and highlight its uniquely low cost (<0.5 USD), speed (sample-to-answer <8 min), multiplexability (12 samples), low volume whole blood requirement (<3 μL), rotation speeds (<4000 rpm) needed for effective measurement as well as the direct finger-to-chip sample loading capability.
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•A 12-sample hematocrit device was developed from polyester-toner materials.•The device can analyze a patient's hematocrit within 8 min from 3 μL of blood.•Cell phone image analysis is used to correctly determine clinical hematocrits.
We describe the first miniaturized device capable of the front-end sample preparation essential for detecting RNA-based infectious agents. The microfluidic device integrates sample purification and ...reverse transcription PCR (RT-PCR) amplification for the identification and detection of influenza A. The device incorporates a chitosan-based RNA binding phase for the completely aqueous isolation of nucleic acids, avoiding the PCR inhibitory effects of guanidine and isopropanol used in silica-based extraction methods. The purified nucleic acids and the reagents needed for single-step RT-PCR amplification are fluidically mobilized simultaneously to a PCR chamber. Utilizing infrared (IR)-mediated heating allowed for a > 5-fold decrease in RT-PCR analysis time compared to a standard thermal cycling protocol used in a conventional thermal cycler. Influenza A virus A/PR/8/34 (H1N1) was used as a simulant in this study for virus-based infectious and biowarfare agents with RNA genomes, and was successfully detected in a mock nasal swab sample at clinically relevant concentrations. Following on-chip purification, a fragment specific to the influenza A nucleoprotein gene was first amplified via RT-PCR amplification using IR-mediated heating to achieve more rapid heating and cooling rates. This was initially accomplished on a two-chip system to optimize the SPE and RT-PCR, and then translated to an integrated SPE-RT-PCR device.
In the last decade, the microfluidic community has witnessed an evolution in fabrication methodologies that deviate from using conventional glass and polymer-based materials. A leading example within ...this group is the print, cut and laminate (PCL) approach, which entails the laser cutting of microfluidic architecture into ink toner-laden polyester sheets, followed by the lamination of these layers for device assembly. Recent success when applying this method to human genetic fingerprinting has highlighted that it is now ripe for the refinements necessary to render it amenable to mass-manufacture. In this communication, we detail those modifications by identifying and implementing a suitable heat-sensitive adhesive (HSA) material to equip the devices with the durability and resilience required for commercialization and fieldwork. Importantly, this augmentation is achieved without sacrificing any of the characteristics which make the PCL approach attractive for prototyping. Exemplary HSA-devices performed DNA extraction, amplification and separation which, when combined, constitute the complete sequence necessary for human profiling and other DNA-based analyses.
The lack of a clear framework identifying data to link ecosystems to analyses of human well-being has been highlighted in numerous studies. To address this issue, we applied a recently developed ...economic theory termed "final" ecosystem goods and services - the biophysical features and qualities that people perceive as being directly related to their well-being. The six-step process presented here enabled us to identify metrics associated with streams that can be used in the analysis of human well-being; we illustrate these steps with data from a regional stream survey. Continued refinement and application of this framework will require ongoing collaboration between natural and social scientists. Framework application could result in more useful and relevant data, leading to more informed decisions in the management of ecosystems.
Chitosan-coated silica particles and chitosan-coated microchannels have been explored as an alternative to a standard silica phase for DNA extraction in a microdevice (Cao, W.; Easley, C. J.; ...Ferrance, J. P.; Landers, J. P. Anal. Chem. 2006, 78 (20), 7222−7228). A method that exploits the use of aqueous buffers for nucleic acid binding to and release from a solid phase is advantageous, avoiding the reagents used for conventional extraction (isopropanol and guanadinium hydrochloride), which are potent PCR inhibitors. The pH-controlled approach, which promotes nucleic acid binding to and release to the chitosan phase based on a change in buffer pH, is exploited here for RNA purification in a microfluidic device. The chitosan phase reproducibly allowed for higher RNA extraction efficiencies under aqueous conditions (71%) compared to that with a silica phase under chaotropic conditions (53%). The effectiveness of the chitosan phase was demonstrated with the successful purification of RNA from the alveolar rhabdomyosarcoma (ARMS) cancer cell line, with 3.5-fold greater extraction efficiencies than obtained when the same sample was purified using a silica phase: the resulting RNA was found to be amplifiable in reverse-transcription PCR. Low-molecular weight chitosan is also a proven inhibitor of RNases, further demonstrating the advantages of chitosan as a solid phase for RNA purification compared to silica. The chitosan phase is, therefore, a superior choice for extraction and purification of RNA in a microfluidic device and is compatible with biological samples found in a clinical or forensic setting.
As COVID-19 transmission control measures are gradually being lifted, a sensitive and rapid diagnostic method for large-scale screening could prove essential for monitoring population infection ...rates. However, many rapid workflows for SARS-CoV-2 detection and diagnosis are not amenable to the analysis of large-volume samples. Previously, our group demonstrated a technique for SARS-CoV-2 nanoparticle-facilitated enrichment and enzymatic lysis from clinical samples in under 10 min. Here, this sample preparation strategy was applied to pooled samples originating from nasopharyngeal (NP) swabs eluted in viral transport medium (VTM) and saliva samples diluted up to 1:100. This preparation method was coupled with conventional RT-PCR on gold-standard instrumentation for proof-of-concept. Additionally, real-time PCR analysis was conducted using an in-house, ultra-rapid real-time microfluidic instrument paired with an experimentally optimized rapid protocol. Following pooling and extraction from clinical samples, average cycle threshold (C
) values from resultant eluates generally increased as the pooling dilution factor increased; further, results from a double-blind study demonstrated 100% concordance with clinical values. In addition, preliminary data obtained from amplification of eluates prepared by this technique and analyzed using our portable, ultra-rapid real-time microfluidic PCR amplification instrument showed progress toward a streamlined method for rapid SARS-CoV-2 analysis from pooled samples.
A glass microdevice has been constructed for the on-line integration of solid-phase extraction (SPE) of DNA and polymerase chain reaction (PCR) on a single chip. The chromatography required for SPE ...in the microfluidic sample preparation device (μSPD) was carried out in a silica bead/sol−gel SPE bed, where the purified DNA was eluted directly into a downstream chamber where conventional thermocycling allowed for PCR amplification of specific DNA target sequences. Through rapid, simple passivation of the PCR chamber with a silanizing reagent, reproducible DNA extraction and amplification was demonstrated from complex biological matrixes in a manner amenable to any research laboratory, using only a syringe pump and a conventional thermocycler. The μSPD allowed for SPE concentration of DNA from 600 nL of blood coupled to subsequent on-chip amplification that yielded a detectable amplicon; this simple device can be applied to a variety of routine genetic analyses without the need for sophisticated instrumentation. In addition, the applicability of these developments to nonconventional thermocycling was demonstrated through the use of noncontact, IR-mediated heating. This was exemplified with the isolation of DNA from an anthrax spore-spiked nasal swab and the subsequent on-chip amplification of target DNA sequences in a total processing time of only 25 min.
Abstract
Introduction
Understanding the changing epidemiology of adults hospitalized with coronavirus disease 2019 (COVID-19) informs research priorities and public health policies.
Methods
Among ...adults (≥18 years) hospitalized with laboratory-confirmed, acute COVID-19 between 11 March 2021, and 31 August 2022 at 21 hospitals in 18 states, those hospitalized during the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron-predominant period (BA.1, BA.2, BA.4/BA.5) were compared to those from earlier Alpha- and Delta-predominant periods. Demographic characteristics, biomarkers within 24 hours of admission, and outcomes, including oxygen support and death, were assessed.
Results
Among 9825 patients, median (interquartile range IQR) age was 60 years (47–72), 47% were women, and 21% non-Hispanic Black. From the Alpha-predominant period (Mar–Jul 2021; N = 1312) to the Omicron BA.4/BA.5 sublineage-predominant period (Jun–Aug 2022; N = 1307): the percentage of patients who had ≥4 categories of underlying medical conditions increased from 11% to 21%; those vaccinated with at least a primary COVID-19 vaccine series increased from 7% to 67%; those ≥75 years old increased from 11% to 33%; those who did not receive any supplemental oxygen increased from 18% to 42%. Median (IQR) highest C-reactive protein and D-dimer concentration decreased from 42.0 mg/L (9.9–122.0) to 11.5 mg/L (2.7–42.8) and 3.1 mcg/mL (0.8–640.0) to 1.0 mcg/mL (0.5–2.2), respectively. In-hospital death peaked at 12% in the Delta-predominant period and declined to 4% during the BA.4/BA.5-predominant period.
Conclusions
Compared to adults hospitalized during early COVID-19 variant periods, those hospitalized during Omicron-variant COVID-19 were older, had multiple co-morbidities, were more likely to be vaccinated, and less likely to experience severe respiratory disease, systemic inflammation, coagulopathy, and death.
Compared to adults hospitalized with Alpha- and Delta-variant COVID-19, those hospitalized later with Omicron-variant COVID-19 were older, had more co-morbidities, were more likely to be vaccinated, and less likely to experience severe respiratory disease, systemic inflammation, coagulopathy, and death.
Graphical Abstract
Graphical Abstract
This graphical abstract is also available at Tidbit: https://tidbitapp.io/tidbits/changing-severity-and-epidemiology-of-adults-hospitalized-with-covid-19-in-the-united-states-after-introduction-of-covid-19-vaccines-march-2021-august-2022-f66f7f84-e1a9-4536-aa93-1e4a50d97d8f
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