Publication detail

Direct magnetic beads-based extraction of microRNA from urine with capillary electrophoretic analysis using fluorescence detection and universal label

ŠMERKOVÁ, K. RÝPAR, T. ADAM, V. VACULOVIČOVÁ, M.

Original Title

Direct magnetic beads-based extraction of microRNA from urine with capillary electrophoretic analysis using fluorescence detection and universal label

Type

journal article in Web of Science

Language

English

Original Abstract

Short non-coding RNAs, specifically microRNAs (miRNAs), are of a great interest due to their presumed function in genome regulation. Moreover, miRNAs are currently perceived as potential biomarkers for numerous diseases; a variety of detection methods and sensing systems have therefore been studied. We present a magnetic-bead-based assay for specific miRNA isolation coupled with sensitive electrophoretic analysis with fluorescence detection. The magnetic separation step involves creating a duplex with targeted miR-141, which is subsequently cleaved from the magnetic bead surface with a specific endonuclease. The duplex is then determined using capillary electrophoresis with laser-induced fluorescence detection in the presence of the fluorescent dye PicoGreen for quantitating double-stranded DNA. The benefits of using microcolumn separation technique coupled with sensitive detection over traditionally used determination by fluorescence spectrometry include the fact that there is no need for a specific pre-labeled fluorescent probe. This significantly simplifies the method and reduces the costs. Cross-reactivity with mismatched oligonucleotides (3 and 5 mismatched bases) and different miRNAs (miR-124 and miR-150) was tested, demonstrating the specificity of the developed method for miRNA-141. This magnetic extraction method was demonstrated for the direct isolation and determination of miR-141 at different concentration levels from urine samples and the achieved nanomolar detection limit.

Keywords

Immobilization; Sample Isolation; miR-141; Endonuclease Cleavage

Authors

ŠMERKOVÁ, K.; RÝPAR, T.; ADAM, V.; VACULOVIČOVÁ, M.

Released

31. 1. 2020

ISBN

1550-7033

Periodical

J BIOMED NANOTECHNOL

Year of study

16

Number

1

State

United States of America

Pages from

76

Pages to

84

Pages count

9

URL

https://doi.org/10.1166/jbn.2020.2872

BibTex

@article{BUT162416,
  author="Kristýna {Šmerková} and Tomáš {Rýpar} and Vojtěch {Adam} and Markéta {Vaculovičová}",
  title="Direct magnetic beads-based extraction of microRNA from urine with capillary electrophoretic analysis using fluorescence detection and universal label",
  journal="J BIOMED NANOTECHNOL",
  year="2020",
  volume="16",
  number="1",
  pages="76--84",
  doi="10.1166/jbn.2020.2872",
  issn="1550-7033",
  url="https://doi.org/10.1166/jbn.2020.2872"
}