Separation behavior of short single‐ and double‐stranded DNA in 1 micron and 100 nm glass channels

Abstract

Micro- and nanofluidic lab-on-chip technology offers the unique capability of high-resolution separation, identification, and manipulation of biomolecules with broad applications in chemistry, biology, and medicine. In this work, we probe the effects of ionic strength on separation of ss- and dsDNA within 1 micron and 100 nm-deep glass channels. Separation behavior of DNA is influenced by a number of parameters, including ionic strength, melting temperature, strand length, strand conformation, and channel size. Specifically, we find a shift in the observed mobility of 10-bp (base pair) dsDNA for different ionic strengths due to changes in kinetic parameters, underlying the importance of these considerations when working with short DNA. For 50-base DNA, the electrophoretic mobility difference between ss- and dsDNA increases as the ionic strength increases due to changes in conformation of the ssDNA. Finally, we find that decreasing channel size decreases the absolute electrophoretic mobility of 10- and 20-bp ss- and dsDNA, due to both hydrodyamic confinement and electric double layer (EDL) interactions. We hypothesize that about 4% mobility reduction is due to hydrodynamic confinement, which is observed at all ionic strengths, and further reduction is due to EDL interactions between the DNA and the channel walls, only observed at low ionic strengths.

Confinement effects on DNA hybridization in electrokinetic micro‐ and nanofluidic systems

“…To further confirm that this confinement effect results from the channel size, we analyze experimental data from previous work using our numerical method. In these experiments, a 16.4 kV/m injection field separates 20 bp DNA oligomers (5’‐AAGAGGAGGGAAGAGGAGGG‐3’ tagged with 6‐fluorescein amidite on the 5’ position and its untagged complement) in a cross‐channel electrophoresis chip . In Fig.…”

“…The simulation solves for the electrophoretic transport of a finite‐sized plug of three reacting species in a straight channel, assuming that the advective velocity equals 0 . This assumption is made because the area‐averaged electroosmotic flow in microchannels falls within 95% of the Helmholtz‐Smoluchowski velocity, allowing us to neglect non‐uniform electroosmotic flow . This numerical simulation utilizes the method of lines technique with upwind finite differencing.…”

Fundamental Studies of Nanofluidics: Nanopores, Nanochannels, and Nanopipets

“… 219 The effects of ionic strength on the separation of single-stranded and double-stranded DNA are probed in glass nanochannels. 220 Electrophoretic mobilities shift at different ionic strengths, and the mobility difference between ss- and dsDNA increases as ionic strength increases. DNA molecules of different lengths are also separated in nanochannels with an asymmetric bipolar pulse.…”

Particle-by-Particle Charge Analysis of DNA-Modified Nanoparticles Using Tunable Resistive Pulse Sensing

“…To build up a true measure of the spread of zeta potential values for a given particle population, the zeta potential of each individual particle has to be measured, and this aspect is challenging, although electrophoretic and electrochemical techniques allow insight to these measurements 29,39 . Electrophoresis studies have demonstrated the ability to separate ssDNA and dsDNA modified particles, and probe the structure of the ssDNA surfaces [40][41][42] . Alternative technologies for monitoring particle-by-particle zeta potentials rely upon particle tracking technologies that monitor the speed of the particles in an applied electric field 43 .…”

“…When using nanoparticle systems a mean population zeta potential will not allow the true measure of the ligand distribution across all of the particles to be interpreted, and in a typical reaction the ligand density would follow a Gaussian distribution. − The spread of the population can have an effect on the reaction kinetics, stability and sensitivity of nanoparticle based assays. − To build up a true measure of the spread of zeta potential values for a given particle population, the zeta potential of each individual particle has to be measured, and this aspect is challenging, although electrophoretic and electrochemical techniques allow insight into these measurements. , Electrophoresis studies have demonstrated the ability to separate ssDNA and dsDNA modified particles, and probe the structure of the ssDNA surfaces. − Alternative technologies for monitoring particle-by-particle zeta potentials rely upon particle tracking technologies that monitor the speed of the particles in an applied electric field …”