Rapid and Sensitive Pathogen Detection by DNA Amplification Using Janus Particle-Enabled Rotational Diffusometry.

Area of Science:

• Biomedical Engineering
• Biosensing Technologies
• Molecular Diagnostics

Background:

• Rapid and sensitive detection of infectious bacteria is crucial for timely medical intervention and infection control.
• Conventional Polymerase Chain Reaction (PCR) methods can be time-consuming for pathogen detection.
• There is a need for faster, more sensitive diagnostic tools for infectious diseases.

Purpose of the Study:

• To develop and validate a novel biosensing method using rotational diffusometry (RD) for the rapid and sensitive detection of *Escherichia coli* (E. coli) DNA.
• To evaluate the performance of RD in terms of sensitivity, detection time, and sample volume compared to conventional PCR.
• To investigate the influence of DNA concentration, amplicon size, and thermal cycles on E. coli detection using RD.

Main Methods:

• Utilized rotational diffusometry (RD), a phenomenon involving Janus particles, to measure changes in fluid viscosity.
• Captured images of microbeads at 10 Hz using a CCD camera and analyzed them with a cross-correlation algorithm.
• Performed gene amplification for specific *E. coli* sequences and assessed detection sensitivity across varying DNA concentrations, amplicon sizes (84, 147, 246 bp), and thermal cycles (10, 20, 30).

Main Results:

• Achieved sensitive detection of *E. coli* DNA at concentrations as low as 50 pg/μL within 30 seconds using 30 thermal cycles.
• Demonstrated detection sensitivity of 0.1 ng/μL and 1 ng/μL DNA with 10 and 20 thermal cycles, respectively, for a 246 bp amplicon.
• The RD method achieved a total turnaround time of less than 60 minutes, significantly faster than conventional PCR.
• Visualized the effects of DNA concentration, amplicon size, and thermal cycles on detection via three distinct maps, highlighting RD's advantages.

Conclusions:

• Rotational diffusometry (RD) provides a rapid, sensitive, and efficient biosensing platform for *E. coli* DNA detection.
• The RD method offers a significant improvement in detection time and turnaround time compared to conventional PCR.
• This technology holds promise for downstream applications in nucleic acid amplification-based pathogen detection and early disease control strategies.