| Enzyme-based colorimetric detection of nucleic acids using peptide nucleic acid-immobilized microwell plates. | |
| | |
MedLine Citation:
|
PMID: 17708676 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
|
The development of label-free or nonlabeling assays for nucleic acids is important in basic biological research and biomedical diagnosis. In this study, we have developed an enzyme-based colorimetric assay for nucleic acids, which combines the robustness of nonlabeling of DNA and RNA samples and the adequate sensitivity of enzymatic reactions. The core of this assay is the use of neutral peptide nucleic acid (PNA) as capture probe and the electrostatic adsorption of horseradish peroxidase (HRP) on hybridized, negatively charged nucleic acids to report the hybridization events, through HRP-catalyzed color reactions of 3,3',5,5'-tetramethylbenzidine and H(2)O(2). The proposed assay has been validated with fully complementary and single base-mismatched DNAs of different chain lengths. The proposed assay has also been validated with total RNA samples extracted from two human cancer cell lines (A 549 lung cancer cell and HeLa cell) for microRNA detection in real samples. Through extensive optimizations of HRP adsorption and nucleic acid hybridization conditions, detection limits of 0.1-0.2 nM for DNA (depending on chain length) and approximately 2 microg of total RNA have been achieved. Surface plasmon resonance spectroscopy has been used to elucidate the HRP adsorption and PNA-nucleic acid hybridizations through real-time measurements and to provide guidance for the development of the colorimetric assay. |
| | |
Authors:
|
Xiaodi Su; Huey Fang Teh; Xiaohui Lieu; Zhiqiang Gao |
Related Documents
:
|
18654466 - Nucleic acid and nucleotide-mediated synthesis of inorganic nanoparticles. 16718586 - Prediction of molecular alignment of nucleic acids in aligned media. 15333916 - In vitro selection from combinatorial nucleic acid libraries. 3927916 - The effect of ring-fluorination on the rate of o-methylation of dihydroxyphenylalanine ... 7666256 - Positional distribution of stearic acid and oleic acid in a triacylglycerol and dietary... 1525246 - Effect of ascorbic acid on neutrophil functions and hypoxanthine/xanthine oxidase-gener... |
Publication Detail:
|
Type: Journal Article; Research Support, Non-U.S. Gov't Date: 2007-08-21 |
Journal Detail:
|
Title: Analytical chemistry Volume: 79 ISSN: 0003-2700 ISO Abbreviation: Anal. Chem. Publication Date: 2007 Sep |
Date Detail:
|
Created Date: 2007-09-14 Completed Date: 2007-12-11 Revised Date: - |
Medline Journal Info:
|
Nlm Unique ID: 0370536 Medline TA: Anal Chem Country: United States |
Other Details:
|
Languages: eng Pagination: 7192-7 Citation Subset: IM |
Affiliation:
|
Institute of Materials Research and Engineering, 3 Research Link, Singapore 117602. xd-su@imre.a-star.edu.sg |
Export Citation:
|
APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
|
Base Sequence Colorimetry / methods* Enzymes, Immobilized / chemistry* Humans Microchemistry Molecular Sequence Data Nucleic Acids / analysis*, chemistry Peptide Nucleic Acids / chemistry* RNA / analysis Sensitivity and Specificity Spectrum Analysis Surface Plasmon Resonance |
| Chemical | |
Reg. No./Substance:
|
0/Enzymes, Immobilized; 0/Nucleic Acids; 0/Peptide Nucleic Acids; 63231-63-0/RNA |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
Previous Document: New class of Ag/AgCl electrodes based on hydrophobic ionic liquid saturated with AgCl.
Next Document: On state-space reduction in multi-strain pathogen models, with an application to antigenic drift in ...