Integrated bacterial cell lysis and DNA extraction using paper-based isotachophoresis

Area of Science:

• Biotechnology and Biomedical Engineering
• Molecular Diagnostics
• Point-of-Care Testing

Background:

• Bacterial infections pose a global health challenge, exacerbated by limited diagnostic capabilities in resource-limited areas.
• Current point-of-care nucleic acid amplification tests are hindered by complex, multi-step traditional sample preparation methods.
• There is a critical need for simplified, rapid, and effective sample preparation techniques for infectious disease diagnostics.

Purpose of the Study:

• To develop and characterize a novel paper-based sample preparation device for bacterial DNA extraction.
• To integrate electrochemical bacterial lysis and isotachophoresis (ITP) for simultaneous DNA purification and concentration.
• To demonstrate the device's efficacy in complex biological matrices for potential point-of-care applications.

Main Methods:

• Development of a paper-based integrated sample preparation module (p-ITPrep).
• Utilized electrokinetic isotachophoresis (ITP) for DNA purification and concentration.
• Integrated electrochemical methods for bacterial cell lysis directly on the paper device.

Main Results:

• The p-ITPrep device achieved an average DNA concentration factor of 12×.
• Successfully extracted bacterial genomic DNA from samples as low as 10^2 CFU/mL *Mycobacterium smegmatis* (Msm).
• Demonstrated efficient DNA extraction from human saliva, serum, and artificial urine within 20 minutes and only 3 user steps.

Conclusions:

• The paper-based ITP sample preparation module (p-ITPrep) offers a significant advancement over conventional nucleic acid extraction protocols.
• This integrated device provides a simple, inexpensive, and versatile solution for bacterial DNA preparation at the point-of-care.
• p-ITPrep has the potential to enhance the accessibility and efficiency of infectious disease diagnostics, especially in low-resource settings.