Strategies for highly sensitive biomarker detection by Rolling Circle Amplification of signals from nucleic acid composed sensors

2011 ◽  
Vol 3 (10) ◽  
pp. 982 ◽  
Author(s):  
Magnus Stougaard ◽  
Sissel Juul ◽  
Felicie F. Andersen ◽  
Birgitta R. Knudsen
Keyword(s):  
Nucleic Acid ◽  
Rolling Circle Amplification ◽  
Biomarker Detection ◽  
Rolling Circle ◽  
Highly Sensitive

scholarly journals Highly sensitive and specific screening of EGFR mutation using a PNA microarray-based fluorometric assay based on rolling circle amplification and graphene oxide

RSC Advances ◽  
2019 ◽  
Vol 9 (66) ◽  
pp. 38298-38308 ◽  
Author(s):  
Xiaojun Xu ◽  
Shu Xing ◽  
Mengjia Xu ◽  
Pan Fu ◽  
Tingting Gao ◽  
...  
Keyword(s):  
Graphene Oxide ◽  
Nucleic Acid ◽  
Peptide Nucleic Acid ◽  
Egfr Mutation ◽  
Rolling Circle Amplification ◽  
Specific Detection ◽  
Fluorometric Method ◽  
Rolling Circle ◽  
Highly Sensitive ◽  
Detection Probe

A facile peptide nucleic acid microarray-based fluorometric method was developed for sensitive and specific detection of EGFR mutation by using rolling circle amplification, graphene oxide, and a fluorescently-labeled detection probe.

The analysis of proteins and small molecules based on sterically tunable nucleic acid hyperbranched rolling circle amplification

2017 ◽  
Vol 91 ◽  
pp. 136-142 ◽  
Author(s):  
Hai Shi ◽  
Xiaoxia Mao ◽  
Xiaoxia Chen ◽  
Zihan Wang ◽  
Keming Wang ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Small Molecules ◽  
Rolling Circle Amplification ◽  
Rolling Circle

Target-responsive dumbbell probe-mediated rolling circle amplification strategy for highly sensitive Hg2+ detection

RSC Advances ◽  
2014 ◽  
Vol 4 (51) ◽  
pp. 27091-27097 ◽  
Author(s):  
Qingwang Xue ◽  
Yanqin Lv ◽  
Yuanfu Zhang ◽  
Shuling Xu ◽  
Qiaoli Yue ◽  
...  
Keyword(s):  
Rolling Circle Amplification ◽  
Selective Detection ◽  
Label Free ◽  
Rolling Circle ◽  
Fluorescent Sensing ◽  
Highly Sensitive ◽  
Mercuric Ions ◽  
Amplification Strategy

A novel label-free amplified fluorescent sensing scheme based on target-responsive dumbbell probe-mediated rolling circle amplification (D-RCA) has been developed for sensitive and selective detection of mercuric ions.

Mitigating the Non-Specific Amplification in RCA: Assessment of the Role of Ligation and Exonuclease Digestion in Circular DNA Preparation

2021 ◽  
Author(s):  
Vandana Kuttappan Nair ◽  
Chandrika Sharma ◽  
Mrittika Sengupta ◽  
Souradyuti Ghosh
Keyword(s):  
Nucleic Acid ◽  
Real Time ◽  
False Positive ◽  
Rolling Circle Amplification ◽  
Circular Dna ◽  
Rolling Circle ◽  
Specific Amplification ◽  
Exonuclease Digestion ◽  
Sticky End

<b>Layman Summary: </b>Rolling circle amplification (RCA) is a popular and extensively used bioanalytical tool. Like any nucleic acid amplifications, non-specific amplification may occur in it and risk generating false positive readouts. The work described in the manuscript investigates non-specific amplification in RCA as a function of ligation and exonuclease digestion assays during the synthesis of circular DNA. In particular, it investigates and compares the role of three different ligation techniques, namely splint-padlock ligation, cohesive end (sticky end ligation), and self-annealing ligation. In addition, it also probes the role of single exonuclease vs dual exonuclease digestions. We employed real time fluorescence to quantify the effect of these factors. Finally, our work hypothesizes the possible origins of non-specific amplification in RCA.

scholarly journals Highly sensitive detection of Staphylococcus aureus by a THz metamaterial biosensor based on gold nanoparticles and rolling circle amplification

RSC Advances ◽  
2020 ◽  
Vol 10 (45) ◽  
pp. 26824-26833 ◽  
Author(s):  
Ke Yang ◽  
Wenjing Yu ◽  
Guorong Huang ◽  
Jie Zhou ◽  
Xiang Yang ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Rolling Circle Amplification ◽  
Rolling Circle ◽  
Hospital Acquired Infections ◽  
Food Borne ◽  
Highly Sensitive ◽  
Food Borne Illness ◽  
Highly Sensitive Detection ◽  
Hospital Acquired ◽  
The Impact

A highly sensitive method for detecting Staphylococcus aureus (S. aureus) is urgently needed to reduce the impact and spread of hospital-acquired infections and food-borne illness.

Molecular Beacon Enables Combination of Highly Processive and Highly Sensitive Rolling Circle Amplification Readouts for Detection of DNA-Modifying Enzymes

Nano LIFE ◽  
2015 ◽  
Vol 05 (02) ◽  
pp. 1541002 ◽  
Author(s):  
Emil L. Kristoffersen ◽  
Maria Gonzalez ◽  
Magnus Stougaard ◽  
Cinzia Tesauro
Keyword(s):  
Real Time ◽  
Dna Cleavage ◽  
Topoisomerase I ◽  
Drug Response ◽  
Molecular Beacon ◽  
Rolling Circle Amplification ◽  
Dna Topoisomerase ◽  
Rolling Circle ◽  
Topoisomerase Ib ◽  
Highly Sensitive

Here we present an optimized readout format for detection of the circularized products from a DNA-based sensor for measurement of DNA-modifying enzymes including DNA Topoisomerase I. The basic design of the DNA-sensor relies on the use of a substrate that can be circularized by the activity of DNA-modifying enzymes like type IB Topoisomerases and subsequently amplified by a rolling circle amplification (RCA) mechanism. The RCA process can be followed in real-time by the addition of a molecular beacon with a fluorophore/quencher pair. Upon hybridization to the amplified product, the fluorophore/quencher pair is separated, giving rise to a fluorescent signal, measurable in pseudo real-time using a qPCR machine or in a fluorimeter. The RCA products in complex with the molecular beacon can subsequently be moved to microscopic slides and analyzed in a fluorescence microscope. We describe the proof of the principle of this molecular beacon-based method combining the qPCR readout format with the standard Rolling circle Enhanced Enzymatic Assay previously reported. Although the qPCR setup is less sensitive, it allows easy, fast, and high-throughput measurement of enzyme activities. Human Topoisomerase IB (TopIB) is a well-known target for the clinically used anticancer drugs of the camptothecin family. The cytotoxic effect of camptothecins correlates directly with the intracellular TopIB activity affecting reversibly the Topoisomerase/DNA cleavage complexes. Therefore, we envisioned that the presented method may find use for the prediction of cellular drug response and for drug screening to discover novel molecules that specifically inhibit TopIB or other DNA-modifying enzymes.

A highly sensitive microsphere-based assay for early detection of Type I diabetes

TECHNOLOGY ◽  
2014 ◽  
Vol 02 (03) ◽  
pp. 200-205 ◽  
Author(s):  
Shyam Sundhar Bale ◽  
Gavrielle Price ◽  
Monica Casali ◽  
Nima Saeidi ◽  
Abhinav Bhushan ◽  
...  
Keyword(s):  
Clinical Symptoms ◽  
Type I Diabetes ◽  
Rolling Circle Amplification ◽  
Limited Range ◽  
Assay Method ◽  
Type I ◽  
Rolling Circle ◽  
Highly Sensitive ◽  
Progression Of Disease ◽  
Time Required

Type 1 Diabetes Mellitus (T1DM) is a T-cell mediated autoimmune disease in which deterioration of insulin producing pancreatic β-cells leads to a state of insulin deficiency. It has been shown that the clinical symptoms of T1DM are preceded by presence of islet cell autoantibodies (ICA) in serum. Radioimmunoassay (RIA) based detection of ICA are the current gold standard for diagnosis of T1DM. While the onset of hyperglycemia is an indicator of onset of T1DM, detection of ICA within the serum is important to differentiate T1DM from ketogenic Type 2 Diabetes (T2D) and Maturity Onset Diabetes of the Young (MODY). Due to their limited range of sensitivity, however, RIA cannot detect ICA at low concentrations in serum which could lead to delay in proper diagnosis and treatment. In addition, the use of radioactive species presents major disadvantages including exposure, waste removal, need for specialized licensed facilities to conduct the tests and the time required for the test (> 24 hours). To overcome these limitations, we have developed a rapid, highly sensitive, fluorescent and microsphere-based assay technique using Rolling Circle Amplification (RCA), to profile T1DM marker antibodies in serum. This assay utilizes the ability of RCA to detect very small amounts of DNA coupled with microsphere-immobilization resulting in an assay which is at least 50 times more sensitive than RIA. Further, this assay method requires very low volume of sample (5 μL), and can be easily adapted to detect other autoantibodies at similar sensitivities while reducing the assay time to ~6 hours. This powerful technique could enable detection of T1DM markers much earlier than current methods and enable earlier intervention to deter the progression of disease. In addition, the modularity of this assay would have implications for enhancing the sensitivities of any standard ELISA technique.

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