scholarly journals Development and evaluation of a rapid DNA preparation method for PCR-based DNA virus detection

2018 ◽  
Vol 6 (4) ◽  
pp. 67
Author(s):  
Tailong Qu ◽  
Dun Zhao ◽  
Runcheng Li ◽  
Meng Ge ◽  
Xinglong Yu

<p>We describe a simple, rapid and resource-saving method of DNA preparation from cultured cells, sera and animal tissues for PCR-based DNA virus detection. The method does not require the proteinase K, ethanol or phenol/chloroform used in conventional methods, and the entire procedure is performed in the same tube, reducing possible cross contamination between samples and the expense of laboratory ware. The protocol utilizes guanidine HCl and sodium dodecyl sulfate successively to lyse cells and dissociate proteins from nucleic acid at high temperature, and precipitates SDS and proteins at low temperature while reducing guanidine HCl concentration sufficiently to permit PCR-based virus detection. This method is extremely low cost, high sensitivity and provides a quick and effective way for clinical and laboratory virus detection, and is especially useful for simultaneous analysis of a large number of samples.</p>

2010 ◽  
Vol 10 (1) ◽  
pp. 80-88
Author(s):  
IGRM Temaja ◽  
G. Suastika ◽  
S.H. Hidayat ◽  
U. Kartosuwondo

Antiserum production and serological assay of Chrysanthemum B Carlavirus (CVB). Virus identification based on spesific reaction between antigen and antibody  in serological assay has been widely applied as a tool for plant virus detection. The aims of this research is  to produce  antiserum of the CVB by  guinea pig immunization using  purified CVB of Cianjur isolate. The antiserum   was used further  for  the  serological test. Serological methods for detection of CVB were I-ELISA, TBIA, western blot and ISEM. The result showed that  guinea pig immunization  using 150 µg of purified virus was able to produce 10.75 ml of antiserum. The antiserum produced had high sensitivity for detection of CVB when examined by I-ELISA and TBIA. Besides its low cost, TBIA allows the samples to be blotted on the nitrocellulose membranes in the field and storage of the membranes for later processing in the laboratory. This feature makes it the metode of  choice for large-scale CVB surveying.


Viruses ◽  
2021 ◽  
Vol 13 (3) ◽  
pp. 466
Author(s):  
Ahmed Mahas ◽  
Norhan Hassan ◽  
Rashid Aman ◽  
Tin Marsic ◽  
Qiaochu Wang ◽  
...  

One important factor for successful disease management is the ability to rapidly and accurately identify the causal agent. Plant viruses cause severe economic losses and pose a serious threat to sustainable agriculture. Therefore, optimization of the speed, sensitivity, feasibility, portability, and accuracy of virus detection is urgently needed. Here, we developed a clustered regularly interspaced short palindromic repeats (CRISPR)-based nucleic acid diagnostic method utilizing the CRISPR–Cas12a system for detecting two geminiviruses, tomato yellow leaf curl virus (TYLCV) and tomato leaf curl New Delhi virus (ToLCNDV), which have single-stranded DNA genomes. Our assay detected TYLCV and ToLCNDV in infected plants with high sensitivity and specificity. Our newly developed assay can be performed in ~1 h and provides easy-to-interpret visual readouts using a simple, low-cost fluorescence visualizer, making it suitable for point-of-use applications.


Author(s):  
Antonia Perju ◽  
Nongnoot Wongkaew

AbstractLateral flow assays (LFAs) are the best-performing and best-known point-of-care tests worldwide. Over the last decade, they have experienced an increasing interest by researchers towards improving their analytical performance while maintaining their robust assay platform. Commercially, visual and optical detection strategies dominate, but it is especially the research on integrating electrochemical (EC) approaches that may have a chance to significantly improve an LFA’s performance that is needed in order to detect analytes reliably at lower concentrations than currently possible. In fact, EC-LFAs offer advantages in terms of quantitative determination, low-cost, high sensitivity, and even simple, label-free strategies. Here, the various configurations of EC-LFAs published are summarized and critically evaluated. In short, most of them rely on applying conventional transducers, e.g., screen-printed electrode, to ensure reliability of the assay, and additional advances are afforded by the beneficial features of nanomaterials. It is predicted that these will be further implemented in EC-LFAs as high-performance transducers. Considering the low cost of point-of-care devices, it becomes even more important to also identify strategies that efficiently integrate nanomaterials into EC-LFAs in a high-throughput manner while maintaining their favorable analytical performance.


2015 ◽  
Vol 2015 ◽  
pp. 1-6 ◽  
Author(s):  
Qiangying Yi ◽  
Gleb B. Sukhorokov ◽  
Jin Ma ◽  
Xiaobo Yang ◽  
Zhongwei Gu

Phase change materials absorb the thermal energy when changing their phases (e.g., solid-to-liquid) at constant temperatures to achieve the latent heat storage. The major drawbacks such as limited thermal conductivity and leakage prevent the PCMs from wide application in desired areas. In this work, an environmentally friendly and low cost approach, layer-by-layer (LbL) assembly technique, was applied to build up ultrathin shells to encapsulate the PCMs and therefore to regulate their changes in volume when the phase change occurs. Generally, the oppositely charged strong polyelectrolytes Poly(diallyldimethylammonium chloride) (PDADMAC) and Poly(4-styrenesulfonic acid) sodium salt (PSS) were employed to fabricate multilayer shells on emulsified octadecane droplets using either bovine serum albumin (BSA) or sodium dodecyl sulfate (SDS) as surfactant. Specifically, using BSA as the surfactant, polyelectrolyte encapsulated octadecane spheres in size of ∼500 nm were obtained, with good shell integrity, high octadecane content (91.3% by mass), and good thermal stability after cycles of thermal treatments.


Micromachines ◽  
2021 ◽  
Vol 12 (6) ◽  
pp. 697
Author(s):  
Siming Lu ◽  
Sha Lin ◽  
Hongrui Zhang ◽  
Liguo Liang ◽  
Shien Shen

Respiratory viral infections threaten human life and inflict an enormous healthcare burden worldwide. Frequent monitoring of viral antibodies and viral load can effectively help to control the spread of the virus and make timely interventions. However, current methods for detecting viral load require dedicated personnel and are time-consuming. Additionally, COVID-19 detection is generally relied on an automated PCR analyzer, which is highly instrument-dependent and expensive. As such, emerging technologies in the development of respiratory viral load assays for point-of-care (POC) testing are urgently needed for viral screening. Recent advances in loop-mediated isothermal amplification (LAMP), biosensors, nanotechnology-based paper strips and microfluidics offer new strategies to develop a rapid, low-cost, and user-friendly respiratory viral monitoring platform. In this review, we summarized the traditional methods in respiratory virus detection and present the state-of-art technologies in the monitoring of respiratory virus at POC.


2021 ◽  
Vol 9 (1) ◽  
Author(s):  
Longfei Li ◽  
Ruirui Shi ◽  
Jianlan Gu ◽  
Yunn Chyn Tung ◽  
Yan Zhou ◽  
...  

AbstractNeurofibrillary tangles (NFTs) made of abnormally hyperphosphorylated tau are a hallmark of Alzheimer’s disease (AD) and related tauopathies. Regional distribution of NFTs is associated with the progression of the disease and has been proposed to be a result of prion-like propagation of misfolded tau. Tau in AD brain is heterogenous and presents in various forms. In the present study, we prepared different tau fractions by sedimentation combined with sarkosyl solubility from AD brains and analyzed their biochemical and pathological properties. We found that tau in oligomeric fraction (O-tau), sarkosyl-insoluble fractions 1 and 2 (SI1-tau and SI2-tau) and monomeric heat-stable fraction (HS-tau) showed differences in truncation, hyperphosphorylation, and resistance to proteinase K. O-tau, SI1-tau, and SI2-tau, but not HS-tau, were hyperphosphorylated at multiple sites and contained SDS- and β-mercaptoethanol–resistant high molecular weight aggregates, which lacked the N-terminal portion of tau. O-tau and SI2-tau displayed more truncation and less hyperphosphorylation than SI1-tau. Resistance to proteinase K was increased from O-tau to SI1-tau to SI2-tau. O-tau and SI1-tau, but not SI2-tau or HS-tau, captured tau from cell lysates and seeded tau aggregation in cultured cells. Heat treatment could not kill the prion-like activity of O-tau to capture normal tau. Hippocampal injection of O-tau into 18-month-old FVB mice induced significant tau aggregation in both ipsilateral and contralateral hippocampi, but SI1-tau only induced tau pathology in the ipsilateral hippocampus, and SI2-tau and HS-tau failed to induce any detectable tau aggregation. These findings suggest that O-tau and SI1-tau have prion-like activities and may serve as seeds to recruit tau and template tau to aggregate, resulting in the propagation of tau pathology. Heterogeneity of tau pathology within AD brain results in different fractions with different biological and prion-like properties, which may pose a major challenge in targeting tau for development of effective therapeutic treatments.


Sensors ◽  
2021 ◽  
Vol 21 (3) ◽  
pp. 942
Author(s):  
Razvan Pascu ◽  
Gheorghe Pristavu ◽  
Gheorghe Brezeanu ◽  
Florin Draghici ◽  
Philippe Godignon ◽  
...  

A SiC Schottky dual-diode temperature-sensing element, suitable for both complementary variation of VF with absolute temperature (CTAT) and differential proportional to absolute temperature (PTAT) sensors, is demonstrated over 60–700 K, currently the widest range reported. The structure’s layout places the two identical diodes in close, symmetrical proximity. A stable and high-barrier Schottky contact based on Ni, annealed at 750 °C, is used. XRD analysis evinced the even distribution of Ni2Si over the entire Schottky contact area. Forward measurements in the 60–700 K range indicate nearly identical characteristics for the dual-diodes, with only minor inhomogeneity. Our parallel diode (p-diode) model is used to parameterize experimental curves and evaluate sensing performances over this far-reaching domain. High sensitivity, upwards of 2.32 mV/K, is obtained, with satisfactory linearity (R2 reaching 99.80%) for the CTAT sensor, even down to 60 K. The PTAT differential version boasts increased linearity, up to 99.95%. The lower sensitivity is, in this case, compensated by using a high-performing, low-cost readout circuit, leading to a peak 14.91 mV/K, without influencing linearity.


2010 ◽  
Vol 9 ◽  
pp. 982-985 ◽  
Author(s):  
Young-Bae Jung ◽  
Jung Han Choi ◽  
Chang Won Jung

2017 ◽  
Vol 9 (6) ◽  
pp. 1031-1037 ◽  
Author(s):  
Jingtao Liu ◽  
Yu Ding ◽  
Lifei Ji ◽  
Xin Zhang ◽  
Fengchun Yang ◽  
...  

Hexavalent chromium (Cr(vi)) is one of the most toxic heavy metal pollutants in groundwater, and thus the detection of Cr(vi) with high sensitivity, accuracy, and simplicity and low cost is of great importance.


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