scholarly journals Nucleic acid amplification by a transparent graphene Visual-PCR chip and a disposable thermocycler

2019 ◽  
Author(s):  
Guozhi Zhu ◽  
Miao Qiao
Keyword(s):  
Nucleic Acids ◽  
Color Change ◽  
Point Of Care ◽  
Low Cost ◽  
Nucleic Acid Amplification ◽  
Driving Voltage ◽  
Conductive Films ◽  
Cooling Fan ◽  
Resource Limited ◽  
Polymerase Chain

ABSTRACTPolymerase chain reaction (PCR) is a method widely used to amplify trace amount of nucleic acids. It needs a process of thermocycling (repeated alternation of temperature). Traditional thermocycler relies on bulk size of metal block to achieve thermocycling, which results in high cost and the lack of portability. Here, a PCR chip made of graphene Transparent Conductive Films (TCFs) was employed. The thermocycling of the chip was fulfilled by a temperature programed microcontroller and a cooling fan under a low driving voltage (12V). A 35 cycles PCR was accomplished within 13 minutes using the chip and the thermocycler. The transparency of the graphene PCR chip enables the PCR reaction to be visually monitored by naked eye for a color change. The PCR chip and the thermocycler have a low cost at $2.5 and $6 respectively, and thus are feasible for Point-of-care testing (POCT) of nucleic acids in a disposable manner. The whole platform makes it possible to perform a low-cost testing of nucleic acids for varieties of purposes outside of laboratories or at resource limited locations.

scholarly journals Nucleic Acids Analytical Methods for Viral Infection Diagnosis: State-of-the-Art and Future Perspectives

Biomolecules ◽  
2021 ◽  
Vol 11 (11) ◽  
pp. 1585
Author(s):  
Emanuele Luigi Sciuto ◽  
Antonio Alessio Leonardi ◽  
Giovanna Calabrese ◽  
Giovanna De Luca ◽  
Maria Anna Coniglio ◽  
...  
Keyword(s):  
Nucleic Acids ◽  
Analytical Methods ◽  
Point Of Care ◽  
Low Cost ◽  
Genetic Material ◽  
Treatment And Prevention ◽  
Polymerase Chain ◽  
Silicon Based ◽  
User Friendly ◽  
Diagnosis Of Infection

The analysis of viral nucleic acids (NA), DNA or RNA, is a crucial issue in the diagnosis of infections and the treatment and prevention of related human diseases. Conventional nucleic acid tests (NATs) require multistep approaches starting from the purification of the pathogen genetic material in biological samples to the end of its detection, basically performed by the consolidated polymerase chain reaction (PCR), by the use of specialized instruments and dedicated laboratories. However, since the current NATs are too constraining and time and cost consuming, the research is evolving towards more integrated, decentralized, user-friendly, and low-cost methods. These will allow the implementation of massive diagnoses addressing the growing demand of fast and accurate viral analysis facing such global alerts as the pandemic of coronavirus disease of the recent period. Silicon-based technology and microfluidics, in this sense, brought an important step up, leading to the introduction of the genetic point-of-care (PoC) systems. This review goes through the evolution of the analytical methods for the viral NA diagnosis of infection diseases, highlighting both advantages and drawbacks of the innovative emerging technologies versus the conventional approaches.

scholarly journals Synthesis, characterisation, and application of chamomile gold nanoparticles in molecular diagnostics: a new component for PCR kits

2019 ◽  
Vol 9 (6) ◽  
pp. 4635-4641 ◽  
Keyword(s):  
Gold Nanoparticles ◽  
Nucleic Acids ◽  
Low Cost ◽  
Gc Content ◽  
Nucleic Acid Amplification ◽  
Chain Reactions ◽  
Polymerase Chain Reactions ◽  
Polymerase Chain ◽  
Successful Amplification

Various strategies have been suggested for successful amplification of the hard-to-amplify nucleic acids such as the inclusion of various chemical or biological materials in the in vitro nucleic acid amplification reactions, particularly polymerase chain reaction (PCR), and adjustment of the cycling programs. Although much efforts have been madefor improvements in the enhancement of polymerase chain reactions of high GC content nucleic acids, still significant challenges remain. In this study, the effects of citrate-coated AuNP and chamomile extract-coated AuNP (p-AuNP) on amplification of three genes with significant different GC percentage were evaluated. Owing to the enormous potential of gold nanoparticles in the enhancement of the PCR reactions, we showed the promising and consistent findings on the application of very dilute biocompatible chamomile-gold nanoparticles as safe and low-cost nanomaterials for molecular amplification of GC-rich DNA samples. We hypothesized that green AuNPs, which have different surface chemistry from cit-AuNPs, not only do not interfere with the PCR reactants but also are capable of enhancing PCR reactions. These results, for the first time, confirm the potential of using the green gold nanoparticles in the heat-assisted enzymatic in vitro reactions, suggesting Chamomile gold nanoparticles as reliable component of any PCR kits.

scholarly journals Validation of a point-of-care rapid diagnostic test for hepatitis C for use in resource-limited settings

2019 ◽  
Vol 11 (4) ◽  
pp. 314-315
Author(s):  
James S Leathers ◽  
Maria Belen Pisano ◽  
Viviana Re ◽  
Gertine van Oord ◽  
Amir Sultan ◽  
...  
Keyword(s):  
Point Of Care ◽  
Low Cost ◽  
Rapid Diagnosis ◽  
Direct Acting Antivirals ◽  
Resource Limited Settings ◽  
Rapid Diagnosis Test ◽  
Resource Limited ◽  
Specificity And Sensitivity ◽  
Hcv Screening ◽  
Direct Acting

Abstract Background Treatment of HCV with direct-acting antivirals has enabled the discussion of HCV eradication worldwide. Envisioning this aim requires implementation of mass screening in resource-limited areas, usually constrained by testing costs. Methods We validated a low-cost, rapid diagnosis test (RDT) for HCV in three different continents in 141 individuals. Results The HCV RDT showed 100% specificity and sensitivity across different samples regardless of genotype or viral load (in samples with such information, 90%). Conclusions The HCV test validated in this study can allow for HCV screening in areas of need when properly used.

scholarly journals Loop Mediated Isothermal Amplification: Principles and Applications in Plant Virology

Plants ◽  
2020 ◽  
Vol 9 (4) ◽  
pp. 461 ◽  
Author(s):  
Stefano Panno ◽  
Slavica Matić ◽  
Antonio Tiberini ◽  
Andrea Giovanni Caruso ◽  
Patrizia Bella ◽  
...  
Keyword(s):  
Plant Protection ◽  
Point Of Care ◽  
Low Cost ◽  
Isothermal Amplification ◽  
Nucleic Acid Amplification ◽  
Plant Virology ◽  
Food Industries ◽  
Loop Mediated Isothermal Amplification ◽  
Application Fields ◽  
Agriculture And Food

In the last decades, the evolution of molecular diagnosis methods has generated different advanced tools, like loop-mediated isothermal amplification (LAMP). Currently, it is a well-established technique, applied in different fields, such as the medicine, agriculture, and food industries, owing to its simplicity, specificity, rapidity, and low-cost efforts. LAMP is a nucleic acid amplification under isothermal conditions, which is highly compatible with point-of-care (POC) analysis and has the potential to improve the diagnosis in plant protection. The great advantages of LAMP have led to several upgrades in order to implement the technique. In this review, the authors provide an overview reporting in detail the different LAMP steps, focusing on designing and main characteristics of the primer set, different methods of result visualization, evolution and different application fields, reporting in detail LAMP application in plant virology, and the main advantages of the use of this technique.

scholarly journals A Rapid, Simple, Inexpensive, and Mobile Colorimetric Assay COVID-19-LAMP for Mass On-Site Screening of COVID-19

2020 ◽  
Vol 21 (15) ◽  
pp. 5380 ◽  
Author(s):  
Franklin Wang-Ngai Chow ◽  
Tony Tat-Yin Chan ◽  
Anthony Raymond Tam ◽  
Suhui Zhao ◽  
Weiming Yao ◽  
...  
Keyword(s):  
Mass Screening ◽  
Color Change ◽  
Colorimetric Assay ◽  
Point Of Care ◽  
Low Cost ◽  
Lamp Assay ◽  
Virus Infections ◽  
Economic Activities ◽  
Amplification Assay ◽  
Rapid Deployment

To control the COVID-19 pandemic and prevent its resurgence in areas preparing for a return of economic activities, a method for a rapid, simple, and inexpensive point-of-care diagnosis and mass screening is urgently needed. We developed and evaluated a one-step colorimetric reverse-transcriptional loop-mediated isothermal amplification assay (COVID-19-LAMP) for detection of SARS-CoV-2, using SARS-CoV-2 isolate and respiratory samples from patients with COVID-19 (n = 223) and other respiratory virus infections (n = 143). The assay involves simple equipment and techniques and low cost, without the need for expensive qPCR machines, and the result, indicated by color change, is easily interpreted by naked eyes. COVID-19-LAMP can detect SARS-CoV-2 RNA with detection limit of 42 copies/reaction. Of 223 respiratory samples positive for SARS-CoV-2 by qRT-PCR, 212 and 219 were positive by COVID-19-LAMP at 60 and 90 min (sensitivities of 95.07% and 98.21%) respectively, with the highest sensitivities among nasopharyngeal swabs (96.88% and 98.96%), compared to sputum/deep throat saliva samples (94.03% and 97.02%), and throat swab samples (93.33% and 98.33%). None of the 143 samples with other respiratory viruses were positive by COVID-19-LAMP, showing 100% specificity. Samples with higher viral load showed shorter detection time, some as early as 30 min. This inexpensive, highly sensitive and specific COVID-19-LAMP assay can be useful for rapid deployment as mobile diagnostic units to resource-limiting areas for point-of-care diagnosis, and for unlimited high-throughput mass screening at borders to reduce cross-regional transmission.

scholarly journals Isothermal Amplification of Nucleic Acids Coupled with Nanotechnology and Microfluidic Platforms for Detecting Antimicrobial Drug Resistance and Beyond

2021 ◽  
Author(s):  
Seyedeh Zahra Alamolhoda ◽  
Nosratollah Zarghami ◽  
Houman Kahroba ◽  
Ahmad Mehdipour ◽  
Mohammad Pourhassan-Moghaddam ◽  
...  
Keyword(s):  
Nucleic Acids ◽  
Point Of Care ◽  
High Sensitivity ◽  
Molecular Techniques ◽  
Isothermal Amplification ◽  
High Tech ◽  
Antimicrobial Drug Resistance ◽  
Resource Limited ◽  
Accessible Point ◽  
Sensitivity Specificity

Antibiotic resistance is one of the serious health-threatening issues globally, the control of which is indispensable for rapid diagnosis and treatment because of the high prevalence and risks of pathogenicity. Traditional and molecular techniques are relatively expensive, complex, and non-portable, requiring facilities, trained personnel, and high-tech laboratories. Widespread and timely-detection is vital to the better crisis management of rapidly spreading infective diseases, especially in low-tech regions and resource-limited settings. Hence, the need for inexpensive, fast, simple, mobile, and accessible point-of-care (POC) diagnostics is highly demanding. Among different biosensing methods, the isothermal amplification of nucleic acids is favorite due to their simplicity, high sensitivity/specificity, rapidity, and portability, all because they require a constant temperature to work. Isothermal amplification methods are utilized for detecting various targets, including DNA, RNA, cells, proteins, small molecules, ions, and viruses. In this paper, we discuss various platforms, applications, and potentials of isothermal amplification techniques for biosensing of antimicrobial resistance. We also evaluate the potential of these methods, coupled with the novel and rapidly-evolving platforms offered by nanotechnology and microfluidic devices.

scholarly journals Point-of-Care Diagnostic Tools for Surveillance of SARS-CoV-2 Infections

2021 ◽  
Vol 9 ◽  
Author(s):  
Dhanasekaran Sakthivel ◽  
David Delgado-Diaz ◽  
Laura McArthur ◽  
William Hopper ◽  
Jack S. Richards ◽  
...  
Keyword(s):  
Large Scale ◽  
Clinical Symptoms ◽  
Point Of Care ◽  
Cost Effective ◽  
Nucleic Acid Amplification ◽  
Diagnostic Tools ◽  
Rt Pcr ◽  
Serological Tests ◽  
Polymerase Chain ◽  
Effective Public Health

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is a recently emerged and highly contagious virus that causes coronavirus disease 2019 (COVID-19). As of August 24, 2021, there were more than 212 million confirmed COVID-19 cases and nearly 4.4 million deaths reported globally. Early diagnosis and isolation of infected individuals remains one of the most effective public health interventions to control SARS-CoV-2 spread and for effective clinical management of COVID-19 cases. Currently, SARS-CoV-2 infection is diagnosed presumptively based on clinical symptoms and confirmed by detecting the viral RNA in respiratory samples using reverse transcription polymerase chain reaction (RT-PCR). Standard RT-PCR protocols are time consuming, expensive, and technically demanding, which makes them a poor choice for large scale and point-of-care screening in resource-poor settings. Recently developed isothermal nucleic acid amplification tests (iNAAT), antigen and/or serological tests are cost-effective to scale COVID-19 testing at the point-of-care (PoC) and for surveillance activities. This review discusses the development of rapid PoC molecular tools for the detection and surveillance of SARS-CoV-2 infections.

scholarly journals Highly performing point-of-care molecular testing for SARS-CoV-2 with RNA extraction and isothermal amplification.

2020 ◽  
Author(s):  
Pierre Garneret ◽  
Etienne Coz ◽  
Elian Martin ◽  
Jean-Claude Manuguerra ◽  
Elodie Brient-Litzler ◽  
...  
Keyword(s):  
Point Of Care ◽  
Low Cost ◽  
Rna Extraction ◽  
Dna Amplification ◽  
Developed Countries ◽  
Temperature Cycling ◽  
Isothermal Amplification ◽  
Nucleic Acid Amplification ◽  
Clinical Samples ◽  
Molecular Testing

In order to respond to the urgent request of massive testing, developed countries perform nucleic acid amplification tests (NAAT) of SARS-CoV-2 in centralized laboratories. Real-time RT - PCR (Reverse transcription - Polymerase Chain Reaction) is used to amplify the viral RNA and enable its detection. Although PCR is 37 years old, it is still considered, without dispute, as the gold standard. PCR is an efficient process, but the complex engineering required for automated RNA extraction and temperature cycling makes it incompatible for use in point of care settings. In the present work, by harnessing progress made in the past two decades in DNA amplification, microfluidics and membrane technologies, we succeeded to create a portable test, in which SARS-CoV-2 RNA is extracted, amplified isothermally by RT - LAMP (Loop-mediated Isothermal Amplification), and detected using intercalating dyes or highly fluorescent probes. Depending on the viral load, the detection takes between twenty minutes and one hour. Using pools of naso-pharyngal clinical samples, we estimated a sensitivity comparable to RT-qPCR (up to a Cycle threshold of 39, equivalent to <0.1 TCID50 per mL) and a 100% specificity, for other human coronaviruses and eight respiratory viruses currently circulating in Europe. We designed and fabricated an easy-to-use portable device called COVIDISC to carry out the test at the point of care. The low cost of the materials along with the absence of complex equipment paves the way towards a large dissemination of this device. The perspective of a reliable SARS-CoV-2 point of care detection, highly performing, that would deliver on-site results in less than one hour opens up a new efficient approach to manage the pandemics.

scholarly journals Inkjet printing of paraffin on paper allows low-cost point-of-care diagnostics for pathogenic fungi

Cellulose ◽  
2020 ◽  
Vol 27 (13) ◽  
pp. 7691-7701 ◽  
Author(s):  
Anusha Prabhu ◽  
M. S. Giri Nandagopal ◽  
Prakash Peralam Yegneswaran ◽  
Hardik Ramesh Singhal ◽  
Naresh Kumar Mani
Keyword(s):  
Filter Paper ◽  
Limit Of Detection ◽  
Point Of Care ◽  
Low Cost ◽  
Pathogenic Fungi ◽  
Sem Analysis ◽  
Qualitative Assessment ◽  
Resource Limited ◽  
Hydrophobic Barrier ◽  
Pore Blockage

Abstract We present a high resolution, ultra-frugal printing of paper microfluidic devices using in-house paraffin formulation on a simple filter paper. The patterns printed using an office inkjet printer formed a selective hydrophobic barrier of 4 ± 1 µm thickness with a hydrophilic channel width of 275 µm. These printed patterns effectively confine common aqueous solutions and solvents, which was verified by solvent compatibility studies. SEM analysis reveals that the solvent confinement is due to pore blockage in the filter paper. The fabricated paper-based device was validated for qualitative assessment of Candida albicans (pathogenic fungi) by using a combination of L-proline β-naphthylamide as the substrate and cinnamaldehyde as an indicator. Our studies reveal that the pathogenic fungi can be detected within 10 min with the limit of detection (LOD) of 0.86 × 106 cfu/mL. Owing to its simplicity, this facile method shows high potential and can be scaled up for developing robust paper-based devices for biomarker detection in resource-limited settings. Graphic abstract

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