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Related Concept Videos

DNA Microarrays02:34

DNA Microarrays

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Microarrays are high-throughput and relatively inexpensive assays that can be automated to analyze large quantities of data at a time. They are used in genome-wide studies to compare gene or protein expression under two varied conditions, such as healthy and diseased states. Microarrays consist of glass or silica slides on which probe molecules are covalently attached through surface functionalization. Most commonly, the slides are prepared through the chemisorption of silanes to silica...
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A Combinatorial Single-cell Approach to Characterize the Molecular and Immunophenotypic Heterogeneity of Human Stem and Progenitor Populations
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Single-cell multiple gene expression analysis based on single-molecule-detection microarray assay for multi-DNA

Lu Li1, Xianwei Wang2, Xiaoli Zhang1

  • 1School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100, China.

Analytica Chimica Acta
|December 7, 2014
PubMed
Summary

We developed a novel single-molecule-detection microarray assay (SMA) for ultra-sensitive, high-selective DNA determination. This assay enables parallel quantification of multiple DNA types in 10 samples with a detection limit as low as 1.3×10(-16) mol L(-1).

Keywords:
Microarray assayQuantum dotsSingle-cell gene expressionSingle-molecule detection

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Area of Science:

  • Biotechnology
  • Molecular Biology
  • Analytical Chemistry

Background:

  • Accurate and sensitive detection of multiple DNA targets is crucial for various biological and diagnostic applications.
  • Existing methods often face limitations in sensitivity, selectivity, or throughput for simultaneous multi-DNA analysis.

Purpose of the Study:

  • To develop and validate an ultra-sensitive and highly selective single-molecule-detection microarray assay (SMA) for multiplexed DNA determination.
  • To demonstrate the applicability of the SMA for single-cell multiple gene expression analysis.

Main Methods:

  • Fabrication of a capture DNA (DNAc) microarray with 10 identical subarrays, each containing 9 spots for detecting 8 types of target DNAs (DNAts) and one blank.
  • Labeling of captured DNAts with quantum dots (QDs) conjugated to detection DNAs (DNAds) with distinct sequences.
  • Acquisition and analysis of fluorescence single-molecule images using a custom-built reader for quantifying DNAts by counting bright dots.

Main Results:

  • The SMA achieved an ultra-low limit of detection of 1.3×10(-16) mol L(-1) for DNA determination.
  • The assay demonstrated high selectivity and enabled parallel quantification of 8 types of DNAts across 10 samples on a single microarray.
  • Successful application of the SMA for simultaneous quantification of cDNAs corresponding to multiple mRNAs in single human breast cancer cells.

Conclusions:

  • The developed SMA is a powerful tool for ultra-sensitive and high-throughput multiplexed DNA detection.
  • The SMA platform shows significant potential for advancing single-cell gene expression profiling and other molecular diagnostic applications.