Rapid and multi-target genotyping of Helicobacter pylori with digital microfluidics

Jinsong Liu, Rongxin Fu, Shuailong Zhang*, Jialu Hou, Hanbin Ma, Siyi Hu, Hang Li, Yanli Zhang, Weian Wang, Bokang Qiao, Baisheng Zang, Xun Min, Feng Zhang, Jie Du*, Shengkai Yan*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

1 Citation (Scopus)

Abstract

Helicobacter pylori (H. pylori) infection correlates closely with gastric diseases such as gastritis, ulcers, and cancer, influencing more than half of the world's population. Establishing a rapid, precise, and automated platform for H. pylori diagnosis is an urgent clinical need and would significantly benefit therapeutic intervention. Recombinase polymerase amplification (RPA)-CRISPR recently emerged as a promising molecular diagnostic assay due to its rapid detection capability, high specificity, and mild reaction conditions. In this work, we adapted the RPA-CRISPR assay on a digital microfluidics (DMF) system for automated H. pylori detection and genotyping. The system can achieve multi-target parallel detection of H. pylori nucleotide conservative genes (ureB) and virulence genes (cagA and vacA) across different samples within 30 min, exhibiting a detection limit of 10 copies/rxn and no false positives. We further conducted tests on 80 clinical saliva samples and compared the results with those derived from real-time quantitative polymerase chain reaction, demonstrating 100% diagnostic sensitivity and specificity for the RPA-CRISPR/DMF method. By automating the assay process on a single chip, the DMF system can significantly reduce the usage of reagents and samples, minimize the cross-contamination effect, and shorten the reaction time, with the additional benefit of losing the chance of experiment failure/inconsistency due to manual operations. The DMF system together with the RPA-CRISPR assay can be used for early detection and genotyping of H. pylori with high sensitivity and specificity, and has the potential to become a universal molecular diagnostic platform.

Original languageEnglish
Article number116282
JournalBiosensors and Bioelectronics
Volume256
DOIs
Publication statusPublished - 15 Jul 2024

Keywords

  • Digital microfluidics
  • Helicobacter pylori
  • RPA-CRISPR
  • Virulence determinant typing

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