DNA Microviscosity Characterization with Particle Diffusometry for Downstream DNA Detection Applications.

Analytical characterization of DNA microviscosity provides critical biophysical insights into nuclear crowding, nucleic acid based pharmaceutical development, and nucleic acid based biosensor device design. However, most viscosity characterization methods require large sample volumes and destructive testing. In contrast, particle diffusometry permits in situ analysis of DNA microviscosity with short measurement times (8 s) using small volumes (<3 μL) which are compatible with DNA preparatory procedures. This unconventional biosensing approach involves measuring the change in sample viscosity using image processing and correlation-based algorithms. Particle diffusometry requires only a fluorescence microscope equipped with a charge-coupled device (CCD) camera and is a nondestructive measurement method. We use particle diffusometry to characterize the effect of DNA topology, length, and concentration on solution viscosity. In addition, we use particle diffusometry to detect the amplification of DNA from Staphylococcus aureus and Klebsiella pneumoniae, two pathogens commonly related to neonatal sepsis. Successful characterization of pathogen amplification with particle diffusometry provides a new opportunity to apply viscosity characterization toward downstream applications in nucleic acid based pathogen detection.