scholarly journals Direct detection of SARS-CoV-2 using non-commercial RT-LAMP reagents on heat-inactivated samples

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Alisa Alekseenko ◽  
Donal Barrett ◽  
Yerma Pareja-Sanchez ◽  
Rebecca J. Howard ◽  
Emilia Strandback ◽  
...  
Keyword(s):  
Large Scale ◽  
Direct Detection ◽  
Dna Polymerases ◽  
Scale Up ◽  
Lamp Assay ◽  
Cost Effective ◽  
Reaction Conditions ◽  
Clinical Patient ◽  
Reverse Transcriptases ◽  
Large Scale Testing

AbstractRT-LAMP detection of SARS-CoV-2 has been shown to be a valuable approach to scale up COVID-19 diagnostics and thus contribute to limiting the spread of the disease. Here we present the optimization of highly cost-effective in-house produced enzymes, and we benchmark their performance against commercial alternatives. We explore the compatibility between multiple DNA polymerases with high strand-displacement activity and thermostable reverse transcriptases required for RT-LAMP. We optimize reaction conditions and demonstrate their applicability using both synthetic RNA and clinical patient samples. Finally, we validate the optimized RT-LAMP assay for the detection of SARS-CoV-2 in unextracted heat-inactivated nasopharyngeal samples from 184 patients. We anticipate that optimized and affordable reagents for RT-LAMP will facilitate the expansion of SARS-CoV-2 testing globally, especially in sites and settings where the need for large scale testing cannot be met by commercial alternatives.

scholarly journals Detection of SARS-CoV-2 using non-commercial RT-LAMP regents and raw samples.

2020 ◽  
Author(s):  
Alisa Alekseenko ◽  
Donal Barrett ◽  
Yerma Pareja-Sanchez ◽  
Rebecca Howard ◽  
Emilia Strandback ◽  
...  
Keyword(s):  
Dna Polymerases ◽  
Scale Up ◽  
Lamp Assay ◽  
Cost Effective ◽  
Economic Resources ◽  
Strand Displacement ◽  
Reaction Conditions ◽  
Clinical Patient ◽  
Displacement Activity ◽  
Reverse Transcriptases

RT-LAMP detection of SARS-CoV-2 has been shown as a valuable approach to scale up COVID-19 diagnostics and thus contribute to limiting the spread of the disease. Here we present the optimization of highly cost-effective in-house produced enzymes, and we benchmark their performance against commercial alternatives. We explore the compatibility between multiple DNA polymerases with high strand-displacement activity and thermostable reverse transcriptases required for RT-LAMP. We optimize reaction conditions and demonstrate their applicability using both synthetic RNA and clinical patient samples. Finally, we validated the optimized RT-LAMP assay for the detection of SARS-CoV-2 in raw nasopharyngeal samples from 184 patients. We anticipate that optimized and affordable reagents for RT-LAMP will facilitate the expansion of SARS-CoV-2 testing globally, especially in sites and settings with limited economic resources.

scholarly journals NaIO4/Br- as a mild system for the oxidation of 1-methyl-anthra-9,10-quinones

2015 ◽  
Author(s):  
Marcin Cybulski ◽  
Adam Formela ◽  
Katarzyna Sidoryk ◽  
Olga Michalak ◽  
Anna Rosa ◽  
...  
Keyword(s):  
High Pressure ◽  
Carboxylic Acids ◽  
Scale Up ◽  
Lithium Bromide ◽  
Cost Effective ◽  
Building Blocks ◽  
Multidrug Resistant ◽  
Resistant Cell ◽  
Direct Oxidation ◽  
Reaction Conditions

One of the anthraquinone classes comprises compounds with a carbonyl group. These natural or synthetic anthraquinones find their application as building blocks in the synthesis of the compounds with a biological activity. Recently, 4-substituted anthra-9,10-quinone-1-carboxylic acids (2) have been used as key intermediates in the synthesis of patented compounds (3) with anticancer activity against multidrug resistant cell lines. Although 2,7-dihydro-3H-dibenz[de,h]cinnolin-3,7-diones (3) were successfully synthetized in a small laboratory scale, several problems were observed during the preparation of their acid intermediates (2) in a multi-gram scale. The known methods for the preparation of 2 are based on the oxidation of the methyl group in anthra-9,10-quinones (1). The most common are: the oxidation with the diluted nitric acid under high pressure in a sealed tube at the temperature of 195-220 oC, the oxidation in nitrobenzene by passing chlorine gas through the reaction mixture at the temperature of 160-170 oC or in a presence of the fuming sulphuric acid. The mentioned methods require aggressive reagents and specific reaction conditions including high pressure and temperature. Thus, there was a need to find a new efficient, cost-effective and reproducible synthetic method of preparation of 2. While searching literature it was found that the direct oxidation of alkylarenes mediated by the sodium periodate/lithium bromide combination produces benzyl acetates throughout benzyl bromides in the acetic acid, or benzylic acids in the diluted inorganic acid. Based on these results we examined a variety of reaction conditions with or without the bromine source and the oxidizing anion. As a result, a novel procedure for the preparation of highly pure 4-substituted anthra-9,10-quinone-1-carboxylic acids (HPLC > 99.5%) using oxidizing anion/ brominating reagent system was developed. It enabled 2 isolation by the simple filtration of the reaction mixture and was applied in the scale-up of 2,7-dihydro-3H-dibenz[de,h]cinnolin-3,7-dione derivatives.

Biological Synthesis of Silver Nanoparticles and their Biomedical Activity: A Review

2021 ◽  
Vol 09 ◽  
Author(s):  
Sarvat Zafar ◽  
Aiman Zafar ◽  
Fakhra Jabeen ◽  
Miad Ali Siddiq
Keyword(s):  
Silver Nanoparticles ◽  
Large Scale ◽  
Scale Up ◽  
Cost Effective ◽  
Biological Properties ◽  
Single Step ◽  
Biological Synthesis ◽  
Nanosized Particles ◽  
Environment Friendly ◽  
Physical And Chemical

: Nanotechnology studies the various phenomena of physio-chemical procedures and biological properties for the generation of nanosized particles, and their rising challenges in the various sectors, like medicine, engineering, agriculture, electronic, and environmental studies. The nanosized particles exhibit good anti-microbial, anti-inflammatory, cytotoxic, drug delivery, anti-parasitic, anti-coagulant and catalytic properties because of their unique dimensions with large surface area, chemical stability and higher binding density for the accumulation of various bio-constituents on their surfaces. Biological approaches for the synthesis of silver nanoparticles (AgNPs) have been reviewed because it is an easy and single-step protocol and a viable substitute for the synthetic chemical-based procedures. Physical and chemical approaches for the production of AgNPs are also mentioned herein. Biological synthesis has drawn attention because it is cost-effective, faster, non-pathogenic, environment-friendly, easy to scale-up for large-scale synthesis, and having no demand for usage of high pressure, energy, temperature, or noxious chemical ingredients, and safe for human therapeutic use. Therefore, the collaboration of nanomaterials with bio-green approaches could extend the utilization of biological and cytological properties compatible with AgNPs. In this perspective, there is an immediate need to develop ecofriendly and biocompatible techniques, which strengthen efficacy against microbes and minimize toxicity for human cells. The present study introduces the biological synthesis of silver nanoparticles, and their potential biomedical applications have also been reviewed.

Measuring Satisfaction With Standard Survey Instruments and Single-Button Responses on Kiosks

2018 ◽  
Vol 62 (1) ◽  
pp. 1429-1433 ◽  
Author(s):  
Laura R. Rabbitt ◽  
Jacob A. Hasselgren ◽  
Cynthia Cook ◽  
Yevgeniy B. Sirotin
Keyword(s):  
User Satisfaction ◽  
Large Scale ◽  
Cost Effective ◽  
Strongly Correlated ◽  
Alternative Technologies ◽  
Large Scale Testing ◽  
System Usability Scale ◽  
Efficiency And Effectiveness ◽  
Survey Instruments ◽  
Cost Effective Alternative

User satisfaction with a technology is an essential usability metric. Unlike efficiency and effectiveness, which are generally recorded during use, satisfaction is often measured subsequently using questionnaires, such as the modified version of the system usability scale (MSUS). This makes satisfaction the most costly usability measure to acquire in large-scale testing. To mitigate this cost, we compared the performance of a four-button kiosk with a standard SUS instrument for measuring satisfaction. Three hundred and fifty four demographically diverse subjects used the kiosk and completed a SUS questionnaire immediately after using one of two different alternative technologies. Kiosk ratings took only 11.43 ( sd = 7.30) seconds on average to collect, much faster than 1200 seconds on average for the SUS. Kiosk ratings and MSUS scores were strongly correlated ( r = 0.62, p < .005), showing the same pattern of differences between the tested technologies. However, the index of dispersion for kiosk ratings was 71.74% larger than for MSUS scores. We conclude that satisfaction kiosks are a cost-effective alternative for measuring satisfaction in usability studies with large sample sizes.

scholarly journals RAPID AND COST-EFFECTIVE PROCESS BASED ON INSECT LARVAE FOR SCALE-UP PRODUCTION OF SARS-COV-2 SPIKE PROTEIN FOR SEROLOGICAL COVID-19 TESTING

2021 ◽  
Author(s):  
Ignacio Smith ◽  
Gregorio Mc Callum ◽  
Adriana Sabljic ◽  
Juan Marfia ◽  
Silvina Bombicino ◽  
...  
Keyword(s):  
Large Scale ◽  
Scale Up ◽  
High Sensitivity ◽  
Cost Effective ◽  
Elisa Method ◽  
Insect Larvae ◽  
S Protein ◽  
Single Antigen ◽  
Antibody Kinetics ◽  
Virus Strains

Serology testing for COVID-19 is important in evaluating active immune response against SARS-CoV-2, studying the antibody kinetics, and monitoring reinfections with genetic variants and new virus strains, in particular, the duration of antibodies in virus-exposed individuals and vaccine-mediated immunity. In this work, recombinant S protein of SARS-CoV-2 was expressed in Rachiplusia nu, an important agronomic plague. One larva produces an amount of S protein sufficient for 150 determinations in the ELISA method herein developed. We established a rapid production process for SARS-CoV-2 S protein that showed immunoreactivity for anti-SARS-CoV-2 antibodies and was used as a single antigen for developing the ELISA method with high sensitivity (96.2%) and specificity (98.8%). Our findings provide an efficient and cost-effective platform for large-scale S protein production, and the scale-up is linear, thus avoiding the use of complex equipment like bioreactors.

scholarly journals Molecular technologies for detection and typing of bluetongue virus

2017 ◽  
Vol 73 (5) ◽  
pp. 259-264
Author(s):  
Wiesław Niedbalski
Keyword(s):  
Large Scale ◽  
Synthesis Method ◽  
Lamp Assay ◽  
Cost Effective ◽  
Laboratory Diagnosis ◽  
Molecular Techniques ◽  
Nucleic Acid Hybridization ◽  
Diagnostic Tools ◽  
Rt Pcr ◽  
Animal Pathogens

Rapid and accurate diagnosis plays an important role in the implementation of effective measures to control the spread of disease. Historically, the laboratory diagnosis and typing of BTV were carried out by various serological and virological methods, including virus neutralization (VN) assay, ELISA, as well as virus isolation (VI) in cell cultures or in embryonated chicken eggs. At present, various molecular techniques to detect BTV genome are increasingly used as primary diagnostic tools for the serotyping and epidemiological investigations of BTV. Initially, the viral RNA was detected by simple nucleic acid hybridization technologies. Then, conventional RT-PCR assays were developed and evaluated for the detection of BTV serotypes based on nucleotide sequences of different genome segments. Although RT-PCR, with its increased sensitivity, has advantages over hybridization, it is almost impossible to quantify accurately by regular and multiplex PCR procedures, and regular PCR may produce false positive results. Over the recent years, a number of real-time RT-PCR (rRT-PCR) methods have been described. The rRT-PCR offers certain advantages over conventional RT-PCR assay, as it is more rapid, sensitive, and can provide quantitative as well as qualitative genetic information. It does not use agarose gel electrophoresis, decreases the risk of contamination because it is run within an enclosed tube, and is suitable for large-scale testing and automation. The target amplicon is usually smaller, reducing the potential for problems caused by target degradation. Loop-mediated isothermal amplification (LAMP), a novel rapid, accurate and cost effective gene amplification method, is an autocycling and strand displacement DNA synthesis method. LAMP assays have been applied as a method of detecting a variety of animal pathogens, including BTV. RT- LAMP assay can be a valuable tool complementing the routine laboratory diagnosis of BTV.

scholarly journals Stormbow: A Cloud-Based Tool for Reads Mapping and Expression Quantification in Large-Scale RNA-Seq Studies

2013 ◽  
Vol 2013 ◽  
pp. 1-8 ◽  
Author(s):  
Shanrong Zhao ◽  
Kurt Prenger ◽  
Lance Smith
Keyword(s):  
Data Analysis ◽  
Large Scale ◽  
Scale Up ◽  
Local Environment ◽  
Transcriptome Profiling ◽  
Cost Effective ◽  
Rna Seq ◽  
Practical Challenge ◽  
Amazon Web Services ◽  
Computational Resources

RNA-Seq is becoming a promising replacement to microarrays in transcriptome profiling and differential gene expression study. Technical improvements have decreased sequencing costs and, as a result, the size and number of RNA-Seq datasets have increased rapidly. However, the increasing volume of data from large-scale RNA-Seq studies poses a practical challenge for data analysis in a local environment. To meet this challenge, we developed Stormbow, a cloud-based software package, to process large volumes of RNA-Seq data in parallel. The performance of Stormbow has been tested by practically applying it to analyse 178 RNA-Seq samples in the cloud. In our test, it took 6 to 8 hours to process an RNA-Seq sample with 100 million reads, and the average cost was $3.50 per sample. Utilizing Amazon Web Services as the infrastructure for Stormbow allows us to easily scale up to handle large datasets with on-demand computational resources. Stormbow is a scalable, cost effective, and open-source based tool for large-scale RNA-Seq data analysis. Stormbow can be freely downloaded and can be used out of box to process Illumina RNA-Seq datasets.

An Efficient Straightforward Synthesis of Antidepressant Drug Moclobemide

2020 ◽  
Vol 5 (2) ◽  
pp. 174-1778
Author(s):  
Santoshkumar M. Potdar ◽  
Krishnakant T. Waghmode
Keyword(s):  
Present Method ◽  
Large Scale ◽  
Antidepressant Drug ◽  
Cost Effective ◽  
Reaction Conditions ◽  
Chlorobenzoic Acid ◽  
Free Condition ◽  
Solid Catalysts ◽  
Synthetic Strategy ◽  
High Yields

Efferent, new and simple synthesis for antidepressant drug moclobemide has been developed via two liner steps. Initially, morpholine treated with 60% aqueous solution of 2-bromoethylamine hydrochloride under solvent and catalyst free condition leads to a key intermediate N-(2-aminoethyl)morpholine, which subsequently treated with p-chlorobenzoic acid in presence of commercially available solid catalysts, afforded moclobemide with good yield. Mild reaction conditions, short reaction time, easy workup, cost-effective, environment friendliness and high yields are attractive advantages of the present method, so our synthetic strategy was applicable to large scale manufacturing of moclobemide every conveniently.

scholarly journals The Synthesis of Mannose-6-Phosphate Using Polyphosphate-Dependent Mannose Kinase

Catalysts ◽  
2019 ◽  
Vol 9 (3) ◽  
pp. 250
Author(s):  
Wenlong Zhu ◽  
Miaomiao Gao ◽  
Biqiang Chen ◽  
Tianwei Tan ◽  
Hui Cao ◽  
...  
Keyword(s):  
Scale Up ◽  
Enzymatic Reaction ◽  
Synthesis Method ◽  
Cost Effective ◽  
Industrial Applications ◽  
Key Factors ◽  
Reaction Conditions ◽  
Mannose 6 Phosphate ◽  
Arthrobacter Species ◽  
Optimal Reaction

Mannose-6-phosphate (M6P) is involved in many metabolic pathways in life, and it has important applications in the treatment of diseases. This study explored a cost-effective enzyme catalytic synthesis method of M6P, using polyphosphate-dependent mannose kinase from Arthrobacter species. This synthesis uses polyphosphate to replace expensive ATP, and it is greener and safer than chemical synthesis. This study investigated the effects of key factors such as metal ions, temperature, and substrate addition on this enzymatic reaction, and improved the conversion efficiency. We moreover take advantage of the response surface method to explore the best catalytic conditions synthetically. The conversion was 99.17% successful under the optimal reaction conditions. After a series of optimizations, we carried out a 200 mL scale-up experiment, which proved that the method has good prospects for industrial applications.

scholarly journals Bioprocessing of Marine Chitinous Wastes for the Production of Bioactive Prodigiosin

Molecules ◽  
2021 ◽  
Vol 26 (11) ◽  
pp. 3138
Author(s):  
Thi Hanh Nguyen ◽  
San-Lang Wang ◽  
Dai Nam Nguyen ◽  
Anh Dzung Nguyen ◽  
Thi Huyen Nguyen ◽  
...  
Keyword(s):  
Large Scale ◽  
Antioxidant Activities ◽  
Scale Up ◽  
Cost Effective ◽  
Culture Broth ◽  
Anticancer Activities ◽  
Bacterial Fermentation ◽  
High Productivity ◽  
Initial Ph ◽  
Shrimp Wastes

Recently, microbial prodigiosin (PG) has received much attention due to its numerous beneficial applications. The aim of this study was to establish the bioprocessing of marine chitinous wastes (MCWs) for the cost-effective preparation of PG. Of the MCWs, demineralized shrimp shell powders (de-SSP) were found to be a potential source of carbon/nitrogen (C/N) for PG production by bacterial fermentation using Serratia marcescens strains. Further, PG scale-up production was investigated in a 15 L bioreactor system, and the highest yield (6200 mg/L) was achieved during fermentation using 5 L of a novel-designed culture broth that included 1.60% C/N sources (a de-SSP/casein ratio of 7/3), 0.02% K2SO4, and 0.05% K2HPO4, with an initial pH of 6–7. Fermentation was conducted in the dark at 27.5 °C for 8.0 h. This study was the first to report on the utilization of shrimp wastes for cost-effective, large-scale (5 L/pilot) PG production with high productivity (6200 mg/L) in a short cultivation time. The combination of 0.02% K2SO4 and 0.05% K2HPO4 was also found to be a novel salt composition that significantly enhanced PG yield. The red compound was purified and confirmed as PG after analyzing its HPLC profile, mass, and UV/vis spectra. The purified PG was then tested for its bioactivities and showed effective anticancer activities, moderated antioxidant activities, and novel anti-NO effects.

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