Bioanalytical preparation is at the core of drug discovery and the clinical decision-making process. This area has seen many technological breakthroughs in instrumentation as well as multiple advances in analytical techniques.
Novel technologies aim to isolate the target component from complex biochemical samples, shorten the time from sample collection to diagnosis, and deliver as-yet undiscovered targets. Experts in sample-preparation technologies for detection, identification, and analysis of biomolecules explored the latest R&D developments as well as ready-to-market technologies at Knowledge Foundation’s “Integrating Sample Preparation” conference held last month.
When cancer cells undergo necrosis, their nuclear DNA, organelles, and other cellular debris end up in the bloodstream. The ability to rapidly detect circulating cell-free cancer DNA would be a major advance in early cancer detection and screening.
As opposed to short pieces of DNA generated during the course of normal apoptosis, DNA particulates from necrotic cancer cells generally have high molecular weight (hmw). Therefore, appearance of hmw-DNA can serve as a biomarker of the cancer development, especially at early stages, when over 90% of cancers could still be successfully treated.
Low levels of hmw-DNA present considerable challenges for assay development. Conventional sample-preparation technologies result in significant losses of hmw-DNA, limiting the use of this important biomarker.
“A seamless sample-to-answer diagnostic system is critical when analyzing low-quantity biomarkers in a high-noise background,” said Michael J. Heller, Ph.D., professor of bioengineering and nanoengineering, University of California, San Diego.
“Dielectrophoresis (DEP) is a technology capable of separating cells and bacteria as well as nanoscale objects, so it works in the correct molecular size range. However, DEP applications were previously limited by the fact that separations could only be carried out under low conductance conditions.
“Blood and other biological fluids, which have relatively high conductance, had to be considerably diluted for DEP separation. Dilution results in loss of already rare entities such as circulating cancer DNA biomarkers.”
DEP induces the motion of particles in the alternating current using electrodes of different sizes or electrodes in asymmetric arrangements. Migration of molecules depends on the dielectric differences between the molecules and the media. As a result, some particles aggregate in the DEP low field regions and some migrate into DEP high field regions.
Dr. Heller’s lab identified a set of conditions within narrow brackets of AC electric voltage and frequency that enabled DEP separation of biological nanoparticles in high ionic strength (high conductance) samples, such as blood, buffy coat blood, plasma, or cerebrospinal fluid.
“Our proof-of-concept experiments were done using platinum microelectrode arrays overcoated with a hydrogel layer. While most DEP electrode devices simply disentegrate under high conductance conditions, our devices held just enough to show DEP separation,” continued Dr. Heller.
Biological Dynamics, a start-up from Dr. Heller’s lab, has subsequently improved the technology, creating electrodes capable of withstanding high ionic dielectrophoresis conditions. The company is perfecting the technology with additional capabilities such as PCR or sequencing directly from the array.