One-pot microplate-based chemiluminescent assay coupled with catalytic hairpin assembly amplification for DNA detection

Nowadays, considerable efforts are focused on advancing DNA detection methods, which are extremely important in clinical diagnostics, pathogen determination, gene therapy, and forensic analysis. A one-pot sensitive microplate-based chemiluminescent assay coupled with catalytic hairpin assembly (CHA)...

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Bibliographic Details
Published in:Analytical and bioanalytical chemistry 2020-08, Vol.412 (21), p.5105-5111
Main Authors: Bodulev, Oleg L., Burkin, Konstantin M., Efremov, Eugene E., Sakharov, Ivan Yu
Format: Article
Language:English
Subjects:
Amplification
Analytical Chemistry
Assaying
Assembly
Biochemistry
Blood plasma
Characterization and Evaluation of Materials
Chemiluminescence
Chemistry
Chemistry and Materials Science
Deoxyribonucleic acid
DNA
DNA - analysis
Euroanalysis XX
Food Science
Forensic science
Format
Gene therapy
Horseradish Peroxidase - chemistry
Laboratory Medicine
Limit of Detection
Luminescence
Monitoring/Environmental Analysis
Nucleic Acid Amplification Techniques - methods
Oligonucleotides
Proof of Concept Study
Research Paper
Selectivity
Sensitivity
Streptavidin
Streptavidin - chemistry
Target detection
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Summary:Nowadays, considerable efforts are focused on advancing DNA detection methods, which are extremely important in clinical diagnostics, pathogen determination, gene therapy, and forensic analysis. A one-pot sensitive microplate-based chemiluminescent assay coupled with catalytic hairpin assembly (CHA) amplification for detection of a 35-mer DNA oligonucleotide was developed. To improve the assay sensitivity, a triple amplification strategy based on the application of CHA (1), streptavidin-polyperoxidase conjugate (Stp-polyHRP) (2), and an enhanced chemiluminescent reaction (3) was used. The one-pot format of the assay, where all steps of the DNA determination are performed in the same well without transfer of samples from one test tube to another, increased its precision. The proposed assay detected the target DNA in the fM range and distinguished the target DNA from related DNAs, demonstrating its high sensitivity and high selectivity. Moreover, the assay was applied successfully for the quantitative determination of the target in spiked samples of human plasma. A microplate format of the assay was convenient for the analysis of a large number of samples. This study provides a prospective tool for DNA detection. Graphical abstract
ISSN:1618-2642
1618-2650
DOI:10.1007/s00216-020-02438-6