Krull, Ulrich J.

Research

The overall objective of Krull’s research program is the investigation of chemically-selective surfaces that are suitable for development of rapid and reversible biosensors, and this focus provides training of students in areas of surface chemistry and instrument design. Recent innovations and ongoing work includes: the design of surfaces that operate in conjunction with microfluidics for nucleic acid biosensing; surfaces for DNA capture in sample pre-concentration strategies; immobilized self-contained DNA constructs that incorporate fluorescence signaling capability to avoid labeling of targets, and the development of chemistry for DNA detection by nanoscale biosensors.

There is tremendous demand for analytical technologies that determine nucleic acids for elucidation of genetic factors of health, the development of new therapies, the detection of pathogens, and the identification of organisms. Current protocols for genotyping and expression analysis generally involve cell lysis and nucleic acid isolation, perhaps followed by one or a series of amplification steps, and finally some separation and analysis steps. Numerous spatially resolved approaches for development of massive arrayed technologies have been introduced, multiplexed bead-based approaches are available for large-scale screening, and rapid quantitative nucleic acid analysis has advanced with the development of Real Time Polymerase Chain Reaction. The availability of technologies for selective and sensitive determination of targeted nucleic acid sequences has become staggering, yet such technologies only superficially address what may be the most significant frontier in bioanalysis – the chemical dynamics within a cell (for example, as associated with mRNA and siRNA). Enter the world of biosensors, where quantitative analysis can be achieved, multiplexed analysis is possible, and real-time reversible selective detection is done without separation and amplification.

The investigation of biosensing motifs at a molecular scale offers direction for the creation of nanoparticle technologies and molecular switches. Furthermore, many biological and clinical questions remain when considering cellular studies, such as the issues of nanoparticle toxicity, the effectiveness of various methods to introduce nanoparticles into a cell, and issues such as the fate (e.g. sequestration, degradation, transport) of nanoparticles within the cellular machinery. The most recent work done by Krull’s team is directed at investigation of the fundamental surface chemistry that will support creation of nanoscale nucleic acid analysis tools. Krull’s team is exploring quantum dots (QDs) decorated with oligonucleotides as the basis for biosensors that operate using fluorescence resonance energy transfer (FRET), where the QD excites a nearby dye that is associated with the formation of double-stranded DNA. Semiconductor nanocrystals, now known as “Quantum Dots”, exhibit bright, narrow, size-tunable emission as a consequence of quantum confinement. They also have a number of other favourable properties, including: broad absorption spectra which can lead to Stokes’ shifts in excess of 100 nm, lifetimes which are typically longer than 10 ns, and often better resistance to photobleaching than many fluorophores. The surface of QDs can be modified to selectively bind target analytes, while other molecules can be added for other functions. QDs luminescing at multiple wavelengths can be excited using a single wavelength at the blue end of the spectrum and by 2-photon excitation, and multiplexing schemes combining FRET with QDs have been characterized. Krull’s group has been credited as the first research team to demonstrate multiplexing based on the selection of a mixture of QDs that have different emissions, where each emission is tuned to excite one of a set of distinctive oligonucleotide probes. These experiments have already demonstrated capability of SNP discrimination at sub-nanomolar detection levels. This technology is now being moved into microfluidics channels to achieve benefit of on-the-fly stringency control and convective mixing, and is being investigated for applications in intra-cellular analytical chemistry.

The intention of Krull’s team is to learn to develop molecular diagnostic tools that may ultimately be applied in tissues and cells. Such nanoscale nucleic acid biosensor technology is the “next generation” diagnostics tool, concurrently allowing imaging, determination of chemical dynamics, and potentially even therapeutic intervention. Investigations of such new optical spectroscopy and imaging modalities, intended for practical applications in the life sciences, offers students a superb multidisciplinary training environment that draws on expertise in physics, chemistry, biology, engineering and medicine, and includes exposure to "bench to bedside" strategies.