scholarly journals Accelerated RNA detection using tandem CRISPR nucleases

Author(s):  
Tina Y. Liu ◽  
Gavin J. Knott ◽  
Dylan C.J. Smock ◽  
John J. Desmarais ◽  
Sungmin Son ◽  
...  

Direct, amplification-free detection of RNA has the potential to transform molecular diagnostics by enabling simple on-site analysis of human or environmental samples. CRISPR-Cas nucleases offer programmable RNA-guided recognition of RNA that triggers cleavage and release of a fluorescent reporter molecule1,2, but long reaction times hamper sensitivity and speed when applied to point-of-care testing. Here we show that unrelated CRISPR nucleases can be deployed in tandem to provide both direct RNA sensing and rapid signal generation, thus enabling robust detection of ~30 RNA copies/microliter in 20 minutes. Combining RNA-guided Cas13 and Csm6 with a chemically stabilized activator creates a one-step assay that detected SARS-CoV-2 RNA from nasopharyngeal samples with PCR-derived Ct values up to 29 in microfluidic chips, using a compact imaging system. This Fast Integrated Nuclease Detection In Tandem (FIND-IT) approach enables direct RNA detection in a format amenable to point-of-care infection diagnosis, as well as to a wide range of other diagnostic or research applications.

2016 ◽  
Vol 04 (03) ◽  
pp. 1640016 ◽  
Author(s):  
Zhenfeng Wang

Microfluidics is a multidisciplinary technology which enables the face-lift crossing a wide range of applications such as life science research, point-of-care diagnostics and personal medicine. Polymer materials, especially thermoplastics, are dominating this emerging market due to the low material cost and the ease of mass production. This paper reviews the major fabrication technologies for making polymer, especially thermoplastic microfluidic chips, such as micro tooling, injection molding, bonding and surface treatment. The paper also summarizes the key challenges in fulfilling the needs of next generation microfluidic products.


2019 ◽  
Vol 16 (2) ◽  
pp. 244-257 ◽  
Author(s):  
Marcus Vinicius Nora de Souza ◽  
Cristiane França da Costa ◽  
Victor Facchinetti ◽  
Claudia Regina Brandão Gomes ◽  
Paula Mázala Pacheco

Background: 1,2,3-triazoles are an important class of organic compounds and because of their aromatic stability, they are not easily reduced, oxidized or hydrolyzed in acidic and basic environments. Moreover, 1,2,3-triazole derivatives are known by their important biological activities and have drawn considerable attention due to their variety of properties. The synthesis of this nucleus, based on the click chemistry concept, through the 1,3-dipolar addition reaction between azides and alkynes is a well-known procedure. This reaction has a wide range of applications, especially on the development of new drugs. Methods: The most prominent eco-friendly methods for the synthesis of triazoles under microwave irradiation published in articles from 2012-2018 were reviewed. Results: In this review, we cover some of the recent eco-friendly CuAAC procedures for the click synthesis of 1,2,3-triazoles with remarks to new and easily recoverable catalysts, such as rhizobial cyclic β-1,2 glucan; WEB (water extract of banana); biosourced cyclosophoraose (CyS); egg shell powder (ESP); cyclodextrin (β- CD); fish bone powder; nanoparticle-based catalyst, among others. Conclusion: These eco-friendly procedures are a useful tool for the synthesis of 1,2,3-triazoles, providing many advantages on the synthesis of this class, such as shorter reaction times, easier work-up and higher yields when compared to classical procedures. Moreover, these methodologies can be applied to the industrial synthesis of drugs and to other areas.


Molecules ◽  
2021 ◽  
Vol 26 (12) ◽  
pp. 3667
Author(s):  
Mashooq A. Bhat ◽  
Ahmed M. Naglah ◽  
Siddique Akber Ansari ◽  
Hanaa M. Al-Tuwajiria ◽  
Abdullah Al-Dhfyan

A ChCl: Gly (DESs) promoted environmentally benign method was developed for the first time using the reaction of aryl aldehydes and dimedone to give excellent yields of xanthene analogues. The major application of this present protocol is the use of green solvent, a wide range of substrate, short reaction times, ease of recovery, the recyclability of the catalyst, high reaction yield, and ChCl: Gly as an alternative catalyst and solvent. In addition to this, all the synthesized compounds were evaluated for their in vitro antimycobacterial activity against M. tuberculosis H37Ra (MTB) and M. bovis BCG strains. The compounds 3d, 3e, 3f, and 3j showed significant antitubercular activity against MTB and M. bovis strains with minimum inhibitory concentration (MIC) values of 2.5−15.10 µg/mL and 0.26–14.92 µg/mL, respectively. The compounds 3e, 3f, and 3j were found to be nontoxic against MCF-7, A549, HCT 116, and THP-1 cell lines. All the prepared compounds were confirmed by 1H NMR and 13C NMR analysis.


2021 ◽  
Vol 10 (12) ◽  
pp. 2627
Author(s):  
Pierre-Edouard Fournier ◽  
Sophie Edouard ◽  
Nathalie Wurtz ◽  
Justine Raclot ◽  
Marion Bechet ◽  
...  

The Méditerranée Infection University Hospital Institute (IHU) is located in a recent building, which includes experts on a wide range of infectious disease. The IHU strategy is to develop innovative tools, including epidemiological monitoring, point-of-care laboratories, and the ability to mass screen the population. In this study, we review the strategy and guidelines proposed by the IHU and its application to the COVID-19 pandemic and summarise the various challenges it raises. Early diagnosis enables contagious patients to be isolated and treatment to be initiated at an early stage to reduce the microbial load and contagiousness. In the context of the COVID-19 pandemic, we had to deal with a shortage of personal protective equipment and reagents and a massive influx of patients. Between 27 January 2020 and 5 January 2021, 434,925 nasopharyngeal samples were tested for the presence of SARS-CoV-2. Of them, 12,055 patients with COVID-19 were followed up in our out-patient clinic, and 1888 patients were hospitalised in the Institute. By constantly adapting our strategy to the ongoing situation, the IHU has succeeded in expanding and upgrading its equipment and improving circuits and flows to better manage infected patients.


Author(s):  
Seyed Ali Tabatabaei ◽  
Mohammad Zabetian Targhi

Isolation of microparticles and biological cells on microfluidic chips has received considerable attention due to their applications in numerous areas such as medical and engineering fields. Microparticles separation is of great importance in bioassays due to the need for smaller sample and device size and lower manufacturing costs. In this study, we first explain the concepts of separation and microfluidic science along with their applications in the medical sciences, and then, a conceptual design of a novel inertial microfluidic system is proposed and analyzed. The PDMS spiral microfluidic device was fabricated, and its effects on the separation of particles with sizes similar to biological particles were experimentally analyzed. This separation technique can be used to separate cancer cells from the normal ones in the blood samples. These components required for testing were selected, assembled, and finally, a very affordable microfluidic kit was provided. Different experiments were designed, and the results were analyzed using appropriate software and methods. Separator system tests with polydisperse hollow glass particles (diameter 2–20 µm), and monodisperse Polystyrene particles (diameter 5 & 15 µm), and the results exhibit an acceptable chip performance with 86% of efficiency for both monodisperse particles and polydisperse particles. The microchannel collects particles with an average diameter of 15.8, 9.4, and 5.9 μm at the proposed reservoirs. This chip can be integrated into a more extensive point-of-care diagnostic system to test blood samples.


Micromachines ◽  
2021 ◽  
Vol 12 (7) ◽  
pp. 826
Author(s):  
Yanting Liu ◽  
Xuming Zhang

This review aims to summarize the recent advances and progress of plasmonic biosensors based on patterned plasmonic nanostructure arrays that are integrated with microfluidic chips for various biomedical detection applications. The plasmonic biosensors have made rapid progress in miniaturization sensors with greatly enhanced performance through the continuous advances in plasmon resonance techniques such as surface plasmon resonance (SPR) and localized SPR (LSPR)-based refractive index sensing, SPR imaging (SPRi), and surface-enhanced Raman scattering (SERS). Meanwhile, microfluidic integration promotes multiplexing opportunities for the plasmonic biosensors in the simultaneous detection of multiple analytes. Particularly, different types of microfluidic-integrated plasmonic biosensor systems based on versatile patterned plasmonic nanostructured arrays were reviewed comprehensively, including their methods and relevant typical works. The microfluidics-based plasmonic biosensors provide a high-throughput platform for the biochemical molecular analysis with the advantages such as ultra-high sensitivity, label-free, and real time performance; thus, they continue to benefit the existing and emerging applications of biomedical studies, chemical analyses, and point-of-care diagnostics.


Polymers ◽  
2021 ◽  
Vol 13 (10) ◽  
pp. 1583
Author(s):  
Natalia Guerrero-Alburquerque ◽  
Shanyu Zhao ◽  
Daniel Rentsch ◽  
Matthias M. Koebel ◽  
Marco Lattuada ◽  
...  

Ureido-functionalized compounds play an indispensable role in important biochemical processes, as well as chemical synthesis and production. Isocyanates, and KOCN in particular, are the preferred reagents for the ureido functionalization of amine-bearing compounds. In this study, we evaluate the potential of urea as a reagent to graft ureido groups onto amines at relatively low temperatures (<100 °C) in aqueous media. Urea is an inexpensive, non-toxic and biocompatible potential alternative to KOCN for ureido functionalization. From as early as 1864, urea was the go-to reagent for polyurea polycondensation, before falling into disuse after the advent of isocyanate chemistry. We systematically re-investigate the advantages and disadvantages of urea for amine transamidation. High ureido-functionalization conversion was obtained for a wide range of substrates, including primary and secondary amines and amino acids. Reaction times are nearly independent of substrate and pH, but excess urea is required for practically feasible reaction rates. Near full conversion of amines into ureido can be achieved within 10 h at 90 °C and within 24 h at 80 °C, and much slower reaction rates were determined at lower temperatures. The importance of the urea/amine ratio and the temperature dependence of the reaction rates indicate that urea decomposition into an isocyanic acid or a carbamate intermediate is the rate-limiting step. The presence of water leads to a modest increase in reaction rates, but the full conversion of amino groups into ureido groups is also possible in the absence of water in neat alcohol, consistent with a reaction mechanism mediated by an isocyanic acid intermediate (where the water assists in the proton transfer). Hence, the reaction with urea avoids the use of toxic isocyanate reagents by in situ generation of the reactive isocyanate intermediate, but the requirement to separate the excess urea from the reaction product remains a major disadvantage.


2021 ◽  
Vol 7 (12) ◽  
pp. eabf4355
Author(s):  
Patrick G. Bissett ◽  
Henry M. Jones ◽  
Russell A. Poldrack ◽  
Gordon D. Logan

The stop-signal paradigm, a primary experimental paradigm for understanding cognitive control and response inhibition, rests upon the theoretical foundation of race models, which assume that a go process races independently against a stop process that occurs after a stop-signal delay (SSD). We show that severe violations of this independence assumption at short SSDs occur systematically across a wide range of conditions, including fast and slow reaction times, auditory and visual stop signals, manual and saccadic responses, and especially in selective stopping. We also reanalyze existing data and show that conclusions can change when short SSDs are excluded. Last, we suggest experimental and analysis techniques to address this violation, and propose adjustments to extant models to accommodate this finding.


2021 ◽  
Vol 25 ◽  
Author(s):  
Dhaval B. Patel ◽  
Jagruti A. Parmar ◽  
Siddharth S. Patel ◽  
Unnati J. Naik ◽  
Hitesh D. Patel

: The synthesis of ester containing heterocyclic compounds via multicomponent reaction is one of the most preferable process in the synthetic organic chemistry and medicinal chemistry. Compounds containing ester linkage have a wide range of biological application in the pharmaceutical field. Therefore, many method have been developed for the synthesis of these type of derivatives. However, some of them are carried out in the presence of toxic solvents and catalysts, with lower yields, longer reaction times, low selectivities and by-products. Thus, the development of new synthetic methods for the ester synthesis is required in the medicinal chemistry. As we know, multicomponent reactions (MCRs) are a powerful tool towards the one-pot ester synthesis, so in this article we have reviewed the recent developments in ester synthesis. This work covers selected explanation of methods via multicomponent reactions to explore the methodological development in ester synthesis.


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