scholarly journals Quantitative and Ultrasensitive In-situ Immunoassay Technology for SARS-CoV-2 Detection in Saliva

2021 ◽  
Author(s):  
Luke Lee ◽  
Fei Liu ◽  
Yuchao Chen ◽  
Gianluca Roma
Keyword(s):  
Detection Sensitivity ◽  
Testing Methods ◽  
Health Crisis ◽  
Viral Antigens ◽  
Primary Care Settings ◽  
Sample Pooling ◽  
Chemiluminescent Immunoassay ◽  
Pooling Strategy ◽  
In Situ Detection

Abstract The COVID-19 pandemic has become an immense global health crisis. However, the lack of efficient and sensitive on-site testing methods limits early detection for timely isolation and intervention. Here, we present a Quantitative and Ultrasensitive in-situ Immunoassay Technology for SARS-CoV-2 detection in saliva (QUIT SARS-CoV-2). Our nanoporous membrane resonator generates a rapid oscillating flow to purify and concentrate SARS-CoV-2 virus in saliva by 40 folds for in-situ detection of viral antigens based on chemiluminescent immunoassay within 20 min. This method achieved a detection sensitivity below 10 0 copies/mL viral load, comparable to the bench-top PCR equipment. The portable QUIT SARS-CoV-2 system, allowing rapid and accurate on-site viral screen with high-throughput sample pooling strategy, can be performed at the primary care settings and substantially improve the detection and prevention of COVID-19.

scholarly journals Direct in situ reverse transcriptase-polymerase chain reaction

2001 ◽  
Vol 281 (2) ◽  
pp. C726-C732 ◽  
Author(s):  
Rajesh Kher ◽  
Robert Bacallao
Keyword(s):  
Reverse Transcriptase ◽  
Restriction Enzymes ◽  
Cellular Level ◽  
Detection Sensitivity ◽  
Chain Reactions ◽  
Polymerase Chain Reactions ◽  
A Cell ◽  
Polymerase Chain ◽  
In Situ Detection

In situ hybridization has been used for localization of specific nucleic acid sequences at the cellular level despite providing relatively low-detection sensitivity. In situ reverse transcriptase-polymerase chain reactions (RT-PCR) enhance sensitivity and thus enable localization of low-abundance mRNA in a cell. However, the available methods are fraught with problems of nonspecific amplifications as a result of mispriming and/or amplification from partially digested residual genomic DNA in tissue. Herein, we demonstrate that nonspecific background amplification can be eliminated by pretreatment of samples with restriction enzymes before DNase I digestion. Primers tagged with a far-red shifted fluorescent dye such as Cy5 in PCR reactions allow identification of target mRNA by fluorescence microscopy. These novel modifications lead to increased specificity and rapid in situ detection of cellular mRNA and thus may be used for pathological diagnosis.

In Situ Detection of Hepatitis C Viral Antigens

2003 ◽  
pp. 249-256
Author(s):  
Regino P. González-Peralta ◽  
Krzysztof Krawczynski
Keyword(s):  
Hepatitis C ◽  
Viral Antigens ◽  
In Situ Detection

scholarly journals High-Sensitivity Raman Gas Probe for In Situ Multi-Component Gas Detection

Sensors ◽  
2021 ◽  
Vol 21 (10) ◽  
pp. 3539
Author(s):  
Jinjia Guo ◽  
Zhao Luo ◽  
Qingsheng Liu ◽  
Dewang Yang ◽  
Hui Dong ◽  
...  
Keyword(s):  
Raman Spectroscopy ◽  
Optical Fibers ◽  
Signal Transmission ◽  
High Sensitivity ◽  
Multiple Reflection ◽  
Detection Sensitivity ◽  
Gas Detection ◽  
In Situ Detection ◽  
Lower Background

Multiple reflection has been proven to be an effective method to enhance the gas detection sensitivity of Raman spectroscopy, while Raman gas probes based on the multiple reflection principle have been rarely reported on. In this paper, a multi-reflection, cavity enhanced Raman spectroscopy (CERS) probe was developed and used for in situ multi-component gas detection. Owing to signal transmission through optical fibers and the miniaturization of multi-reflection cavity, the CERS probe exhibited the advantages of in situ detection and higher detection sensitivity. Compared with the conventional, backscattering Raman layout, the CERS probe showed a better performance for the detection of weak signals with a relatively lower background. According to the 3σ criteria, the detection limits of this CERS probe for methane, hydrogen, carbon dioxide and water vapor are calculated to be 44.5 ppm, 192.9 ppm, 317.5 ppm and 0.67%, respectively. The results presented the development of this CERS probe as having great potential to provide a new method for industrial, multi-component online gas detection.

Rapid quantification and in situ detection of nitrifying bacteria in biofilms by monoclonal antibody method

2000 ◽  
Vol 41 (4-5) ◽  
pp. 301-308 ◽  
Author(s):  
N. Noda ◽  
H. Ikuta ◽  
Y. Ebie ◽  
A. Hirata ◽  
S. Tsuneda ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Fluorescent Antibody ◽  
Nitrifying Bacteria ◽  
Fluorescent Antibody Technique ◽  
Antibody Technique ◽  
Antibody Method ◽  
In Situ Detection ◽  
Rapid Quantification

Fluorescent antibody technique by the monoclonal antibody method is very useful and helpful for the rapid quantification and in situ detection of the specific bacteria like nitrifiers in a mixed baxterial habitat such as a biofilm. In this study, twelve monoclonal antibodies against Nitrosomonas europaea (IFO14298) and sixteen against Nitrobacter winogradskyi (IFO14297) were raised from splenocytes of mice (BALB/c). It was found that these antibodies exhibited little cross reactivity against various kinds of heterotrophic bacteria. The direct cell count method using monoclonal antibodies could exactly detect and rapidly quantify N. europaea and N. winogradskyi. Moreover, the distribution of N. europaea and N. winogradskyi in a biofilm could be examined by in situ fluorescent antibody technique. It was shown that most of N. winogradskyi existed near the surface part and most of N. europaea existed at the inner part of the polyethylene glycol (PEG) gel pellet, which had entrapped activated sludge and used in a landfill leachate treatment reactor. It was suggested that this monoclonal antibody method was utilized for estimating and controlling the population of nitrifying bacteria as a quick and favorable tool.

scholarly journals Single Step In Situ Detection of Surface Protein and MicroRNA in Clustered Extracellular Vesicles Using Flow Cytometry

2021 ◽  
Vol 10 (2) ◽  
pp. 319
Author(s):  
Hee Cheol Yang ◽  
Won Jong Rhee
Keyword(s):  
Extracellular Vesicles ◽  
Liquid Biopsy ◽  
Molecular Beacon ◽  
Disease Diagnosis ◽  
Single Step ◽  
Flow Cytometer ◽  
Surface Proteins ◽  
Biomarker Detection ◽  
In Situ Detection

Because cancers are heterogeneous, it is evident that multiplexed detection is required to achieve disease diagnosis with high accuracy and specificity. Extracellular vesicles (EVs) have been a subject of great interest as sources of novel biomarkers for cancer liquid biopsy. However, EVs are nano-sized particles that are difficult to handle; thus, it is necessary to develop a method that enables efficient and straightforward EV biomarker detection. In the present study, we developed a method for single step in situ detection of EV surface proteins and inner miRNAs simultaneously using a flow cytometer. CD63 antibody and molecular beacon-21 were investigated for multiplexed biomarker detection in normal and cancer EVs. A phospholipid-polymer-phospholipid conjugate was introduced to induce clustering of the EVs analyzed using nanoparticle tracking analysis, which enhanced the detection signals. As a result, the method could detect and distinguish cancer cell-derived EVs using a flow cytometer. Thus, single step in situ detection of multiple EV biomarkers using a flow cytometer can be applied as a simple, labor- and time-saving, non-invasive liquid biopsy for the diagnosis of various diseases, including cancer.

scholarly journals Sample-Pooling Strategy for SARS-CoV-2 Detection among Students and Staff of the University of Sannio

Diagnostics ◽  
2021 ◽  
Vol 11 (7) ◽  
pp. 1166
Author(s):  
Immacolata Polvere ◽  
Elena Silvestri ◽  
Lina Sabatino ◽  
Antonia Giacco ◽  
Stefania Iervolino ◽  
...  
Keyword(s):  
Quantitative Polymerase Chain Reaction ◽  
Virus Detection ◽  
Testing Procedures ◽  
Stem Infection ◽  
In Vitro Diagnostic ◽  
Sample Pooling ◽  
Pooling Strategy ◽  
Asymptomatic Individuals ◽  
The University

Since the beginning of the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) pandemic, it has been clear that testing large groups of the population was the key to stem infection and prevent the effects of the coronavirus disease of 2019, mostly among sensitive patients. On the other hand, time and cost-sustainability of virus detection by molecular analysis such as reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) may be a major issue if testing is extended to large communities, mainly asymptomatic large communities. In this context, sample-pooling and test grouping could offer an effective solution. Here we report the screening on 1195 oral-nasopharyngeal swabs collected from students and staff of the Università degli Studi del Sannio (University of Sannio, Benevento, Campania, Italy) and analyzed by an in-house developed multiplex RT-qPCR for SARS-CoV-2 detection through a simple monodimensional sample pooling strategy. Overall, 400 distinct pools were generated and, within 24 h after swab collection, five positive samples were identified. Out of them, four were confirmed by using a commercially available kit suitable for in vitro diagnostic use (IVD). High accuracy, sensitivity and specificity were also determined by comparing our results with a reference IVD assay for all deconvoluted samples. Overall, we conducted 463 analyses instead of 1195, reducing testing resources by more than 60% without lengthening diagnosis time and without significant losses in sensitivity, suggesting that our strategy was successful in recognizing positive cases in a community of asymptomatic individuals with minor requirements of reagents and time when compared to normal testing procedures.

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