scholarly journals Immunomagnetic Separation Improves the Detection of Mycobacteria by Paper-Based Lateral and Vertical Flow Immunochromatographic Assays

Sensors ◽  
2021 ◽  
Vol 21 (18) ◽  
pp. 5992
Author(s):  
Alejandra Ben Aissa ◽  
Barbara Araújo ◽  
Esther Julián ◽  
Maria Valnice Boldrin Zanoni ◽  
María Isabel Pividori
Keyword(s):  
Nucleic Acid ◽  
Magnetic Particles ◽  
Immunomagnetic Separation ◽  
Lateral Flow ◽  
Rapid Screening ◽  
Rrna Gene ◽  
Mycobacterium Fortuitum ◽  
Vertical Flow ◽  
Immunosuppressed Patients ◽  
Generating System

This work addresses a method that combines immunomagnetic separation (IMS) and paper-based nucleic acid immunochromatographic assay for the sensitive detection of Mycolicibacterium fortuitum (basonym Mycobacterium fortuitum) In particular, the preconcentration of the bacteria was achieved by using magnetic particles modified with an antibody specific towards mycobacteria. Following the IMS, the bacteria were lysed, and the genome was amplified by double-tagging PCR, using a set of primers specific for the 16S rRNA gene for Mycobacterium. During the amplification, the amplicons were labeled with biotin and digoxigenin tags. Moreover, a comparative study of paper-based immunochromatographic platforms, relying on vertical and lateral flow and on the use of streptavidin gold nanoparticles as a signal generating system, was also performed. The visual readout was achieved when the gold-modified amplicons were captured by the anti-DIG antibody in the test line. The analytical performance of both methods, nucleic acid vertical flow (NAVF) and nucleic acid lateral flow (NALF), is also discussed. Although NALF showed lower limit of detections (LODs), both NALF and NAVF combined with IMS were able to detect the required LOD in hemodialysis water, becoming two promising and useful techniques for the rapid screening of water supplies in hemodialysis centers, to prevent the exposure of immunosuppressed patients to contaminated sources.

Gold-Aptamer-Nanoconstructs Engineered to Detect Conserved Enteroviral Nucleic Acid Sequences

2019 ◽  
Author(s):  
Veeren Chauhan ◽  
Mohamed M Elsutohy ◽  
C Patrick McClure ◽  
Will Irving ◽  
Neil Roddis ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
In Silico ◽  
Point Of Care ◽  
Lateral Flow ◽  
Nucleic Acid Sequence ◽  
In Silico Screening ◽  
Lateral Flow Assays ◽  
Life Threatening ◽  
Software And Hardware ◽  
Nucleic Acid Sequences

<p>Enteroviruses are a ubiquitous mammalian pathogen that can produce mild to life-threatening disease. Bearing this in mind, we have developed a rapid, accurate and economical point-of-care biosensor that can detect a nucleic acid sequences conserved amongst 96% of all known enteroviruses. The biosensor harnesses the physicochemical properties of gold nanoparticles and aptamers to provide colourimetric, spectroscopic and lateral flow-based identification of an exclusive enteroviral RNA sequence (23 bases), which was identified through in silico screening. Aptamers were designed to demonstrate specific complementarity towards the target enteroviral RNA to produce aggregated gold-aptamer nanoconstructs. Conserved target enteroviral nucleic acid sequence (≥ 1x10<sup>-7</sup> M, ≥1.4×10<sup>-14</sup> g/mL), initiates gold-aptamer-nanoconstructs disaggregation and a signal transduction mechanism, producing a colourimetric and spectroscopic blueshift (544 nm (purple) > 524 nm (red)). Furthermore, lateral-flow-assays that utilise gold-aptamer-nanoconstructs were unaffected by contaminating human genomic DNA, demonstrated rapid detection of conserved target enteroviral nucleic acid sequence (< 60 s) and could be interpreted with a bespoke software and hardware electronic interface. We anticipate our methodology will translate in-silico screening of nucleic acid databases to a tangible enteroviral desktop detector, which could be readily translated to related organisms. This will pave-the-way forward in the clinical evaluation of disease and complement existing strategies at overcoming antimicrobial resistance.</p>

scholarly journals Limited-Resource Preparable Chitosan Magnetic Particles for Extracting Amplification-Ready Zeptomole-Range Nucleic Acid from Complex Biofluid

2021 ◽  
Author(s):  
Sayantan Tripathy ◽  
Arunansu Talukdar ◽  
Goutam Pramanik ◽  
P. V. Rajesh ◽  
Souradyuti Ghosh
Keyword(s):  
Nucleic Acid ◽  
Real Time ◽  
Real Time Pcr ◽  
Magnetic Particles ◽  
Limited Resource ◽  
Nucleic Acid Amplification ◽  
Acid Extraction ◽  
Analytical Sensitivity ◽  
Nucleic Acid Extraction ◽  
Spin Column

<b>Layman Summary: </b>Nucleic acid extraction is a key prerequisite for any nucleic acid amplification test (NAAT) or isothermal NAAT (iNAAT) based molecular diagnosis assays.<b> </b>Existing methods utilizes spin column system for nucleic acid extraction which are unsuitable for limited resource settings. Our work explores two methods for chitosan coated magnetic particle preparation that can be executed within 6 h from commonly available chemicals with nothing but a magnetic stirrer and water bath and doable by a minimally trained person. We will also investigated the compatibility of the extracted nucleic acid with downstream NAATs such as real time LAMP, colorimetric LAMP, and real time PCR. In the process, we established the analytical sensitivity of the overall method.<div><br><div><b>Characterization methods</b>: SEM, XRD, EDX, FT-IR</div><div><br></div><div><b>Bioanalytical methods:</b> Real time LAMP, Colorimetric LAMP, Real time PCR</div></div>

scholarly journals Development and Performance verification of colloidal gold labeled SARS-CoV-2 antigen detection method for routine popular screening of COVID-19 with clinical samples in Poland and China

2021 ◽  
Author(s):  
Chuanxiang Guo ◽  
Li Yao ◽  
Fengling Chen ◽  
Chao Zhang ◽  
Wei Chen
Keyword(s):  
Predictive Value ◽  
Colloidal Gold ◽  
Detection Method ◽  
Age Distribution ◽  
Antigen Detection ◽  
Lateral Flow ◽  
Rapid Screening ◽  
Clinical Samples ◽  
Lateral Flow Strip ◽  
And Performance

In this research, we have constructed and optimized the colloidal gold labeled lateral flow strip (LFS) for rapid detection of antigen of SARS-CoV-2 and rapid screening of COVID-19. Based on the constructed and optimized colloidal gold lateral flow strip, the parameters of the LFS have been well evaluated with the clinical samples in the professional labs. The screening performance have also been evaluated from the aspects including the CT values, age distribution and onset of symptoms. Finally, based on the detection results of 420 clinical samples, the LFS can achieve the screening of COVID-19 with the positive percentage agreement (PPA, sensitivity), negative percent agreement (NPA, specificity), the positive predictive value (PPV) and the negative predictive value (NPV) of 96.8%, 100%, 100% and 96.6%, respectively, indicating the powerful potential for practical screening applications in pandemic control. Of great significance, this developed SARS-CoV-2 antigen detection method has also been successfully utilized for screening of delta-variant of SARS-CoV-2.

scholarly journals Study on rapid screening technology of antisense oligonucleotides for SARS-CoV-2

2021 ◽  
Author(s):  
Huawei Shen
Keyword(s):  
Nucleic Acid ◽  
Antisense Oligonucleotides ◽  
Structural Characteristics ◽  
Inhibitory Effect ◽  
Rapid Screening ◽  
Short Term ◽  
Physical Isolation ◽  
The World ◽  
Actual Use ◽  
Screening Technology

Abstract According to the structural characteristics of SARS-CoV-2 nucleic acid,SARS-CoV-2 genomes were virtually segmented.After comparing with human genome,44707 SARS-CoV-2 genomes and 26 primates' genomes, 19 antisense oligonucleotides sequences were selected.Experimental results show that the combined inhibitory effect reaches 100% . Significance statement：Currently, effective methods of controlling the spread of SARS-CoV-2 are very limited, only physical isolation and vaccination.Although physical isolation can achieve some short-term results, but can not fundamentally solve the problem of epidemic spread.The time between the development of a vaccine and its actual use is very long.It often happens that the vaccine is developed successfully, but the epidemic situation is beyond control.If a mutant strain emerges, it will take longer.I hope that the rapid screening technology of SARS-CoV-2 antisense oligonucleotides will light up new hope for the future of mankind and contribute oriental wisdom to the world.

scholarly journals Glycerol Reduces Cross Hybridization on Nitrocellulose Membrane

2020 ◽  
Vol 38 (3) ◽  
pp. 199
Author(s):  
Narendra Yoga Hendarta ◽  
Abu Tholib Aman ◽  
Asmarani Kusumawati ◽  
Tri Wibawa
Keyword(s):  
Nucleic Acid ◽  
Point Of Care ◽  
Gc Content ◽  
Lateral Flow ◽  
Hybridization Signal ◽  
Nitrocellulose Membrane ◽  
High Concentration ◽  
Cross Hybridization ◽  
High Stringency ◽  
Stringency Condition

Lateral flow assay (LFD) based nucleic acid lateral flow (NALF)  method has been developed recently. The method met point of care testing (POCT) as simple and rapid procedures, less equipment, and can be performance by less skilled technician. NALF based on nucleic acid hybridizationis  more economical then immunochromatography assay which use antibody-antigen recognition. Cross hybridization has issued while used to differentiate organism with high GC content and high homology as high similarity genome. Some techniques has applied to give high stringency condition avoid cross hybridization reaction but need more procedure to apply. We found glycerol applied to buffer assay could reduce cross hybridization on nitrocellulose membrane. The study used 2 kinds of high stringency buffer as PBS and SSC bases and high concentration of ssDNA amplicon as sample. Without glycerol ingredient gave cross hybridization signal on test line. But used glycerol could reduce those even omitted with PBS based buffer assay. Beside those, glycerol could significantly increased hybridization signal in SSC based buffer assay (p<0.05).

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