Display Settings:

Format

Send to:

Choose Destination
Biosens Bioelectron. 2013 Apr 15;42:503-11. doi: 10.1016/j.bios.2012.10.013. Epub 2012 Oct 22.

A novel bisulfite-microfluidic temperature gradient capillary electrophoresis platform for highly sensitive detection of gene promoter methylation.

Author information

  • 1Department of Cell Biology, Key Laboratory of Cell Biology, Ministry of Public Health, and Key Laboratory of Medical Cell Biology, Ministry of Education, China Medical University, 92 Beier Road, Heping District, Shenyang, Liaoning 110001, PR China.

Abstract

The hypermethylated tumor suppressor gene promoters are widely recognized as promising markers for cancer screening and ideal targets for cancer therapy, however, a major obstacle in their clinical study is highly sensitive screening. To address this limitation, we developed a novel bisulfite-microfluidic temperature gradient capillary electrophoresis (bisulfite-μTGCE) platform for gene methylation analysis by combining bisulfite treatment and slantwise radiative heating system-based μTGCE. Bisulfite-treated genomic DNA (gDNA) was amplified with universal primers for both methylated and unmethylated sequences, and introduced into glass microfluidic chip to perform electrophorectic separation under a continuous temperature gradient based on the formation of heteroduplexes. Eight CDKN2A promoter model fragments with different number and location of methylation sites were prepared and successfully analyzed according to their electrophoretic peak patterns, with high stability, picoliter-scale sample consumption, and significantly increased detection speed. The bisulfite-μTGCE could detect methylated gDNA with a detection limit of 7.5pg, and could distinguish as low as 0.1% methylation level in CDKN2A in an unmethylated background. Detection of seven colorectal cancer (CRC) cell lines with known and unknown methylation statuses of CDKN2A promoter and 20 tumor tissues derived from CRC patients demonstrated the capability of detecting hypermethylation in real-world samples. The wider adaptation of this platform was further supported by the detection of the CDKN2A and MLH1 promoters' methylation statuses in combination. This highly sensitive, fast, and low-consumption platform for methylation detection shows great potential for future clinical applications.

Copyright © 2012 Elsevier B.V. All rights reserved.

PMID:
23246658
[PubMed - indexed for MEDLINE]
PubMed Commons home

PubMed Commons

0 comments
How to join PubMed Commons

    Supplemental Content

    Full text links

    Icon for Elsevier Science
    Loading ...
    Write to the Help Desk