Sally A. Peyman

Diamagnetic repulsion—A versatile tool for label-free particle handling in microfluidic devices

We report the exploration of diamagnetic repulsion forces for the selective manipulation of microparticles inside microfluidic devices. Diamagnetic materials such as polymers are repelled from magnetic fields, an effect greatly enhanced by suspending a diamagnetic object in a paramagnetic Mn(2+) solution. The versatility of diamagnetic repulsion is demonstrated for the trapping, focussing and deflection of polystyrene particles for three example applications. Firstly, magnet pairs with unlike poles facing each other were arranged along a microcapillary to trap plugs of differently functionalised particles for a simultaneous surface-based assay in which biotin was selectively bound to a plug of streptavidin coated particles utilising only 22nL of reagent. Secondly, by slightly modifying the magnetic field design, the rapid focussing of particles into a narrow central stream at a flow rate of 650microms(-1) was accomplished for particle pre-concentration. In a third application, 5 and 10microm polystyrene particles were separated from each other in continuous flow by passing the particle mixture through a microfluidic chamber with a perpendicular magnetic field, a method termed diamagnetophoresis. The separation was investigated between flow rates of 20-100microL h(-1), with full resolution of the particle populations being achieved at 20microL h(-1). These experiments show the potential of diamagnetic repulsion for simple, label-free manipulation of particles and other diamagnetic objects such as cells for a range of bioanalytical techniques.

Rapid on-chip multi-step (bio)chemical procedures in continuous flow : manoeuvring particles through co-laminar reagent streams

We introduce a novel and extremely versatile microfluidic platform in which tedious multi-step biochemical processes can be performed in continuous flow within a fraction of the time required for conventional methods.

Expanding 3D geometry for enhanced on-chip microbubble production and single step formation of liposome modified microbubbles

Micron sized, lipid stabilized bubbles of gas are of interest as contrast agents for ultra-sound (US) imaging and increasingly as delivery vehicles for targeted, triggered, therapeutic delivery. Microfluidics provides a reproducible means for microbubble production and surface functionalisation. In this study, microbubbles are generated on chip using flow-focussing microfluidic devices that combine streams of gas and liquid through a nozzle a few microns wide and then subjecting the two phases to a downstream pressure drop. While microfluidics has successfully demonstrated the generation of monodisperse bubble populations, these approaches inherently produce low bubble counts. We introduce a new micro-spray flow regime that generates consistently high bubble concentrations that are more clinically relevant compared to traditional monodisperse bubble populations. Final bubble concentrations produced by the micro-spray regime were up to 10(10) bubbles mL(-1). The technique is shown to be highly reproducible and by using multiplexed chip arrays, the time taken to produce one millilitre of sample containing 10(10) bubbles mL(-1) was ∼10 min. Further, we also demonstrate that it is possible to attach liposomes, loaded with quantum dots (QDs) or fluorescein, in a single step during MBs formation.

Nanomechanics of Lipid Encapsulated Microbubbles with Functional Coatings

Microbubbles (MBs) are increasingly being proposed as delivery vehicles for targeted therapeutics, as well as being contrast agents for ultrasound imaging. MBs formed with a lipid shell are promising candidates due to their biocompatibility and the opportunity for surface functionalization, both for specific targeting of tissues and as a means to tune their mechanical response for localized ultrasound induced destruction in vivo. Herein, we acquired force-deformation data on coated lipid MBs using tip-less microcantilevers in an atomic force microscope. Model lipid MBs were designed to test the effects of adding a functional coating on the outside of the lipid leaflet, including a protein coat (streptavidin) or the addition of quantum dots (Q-dots) as optical reporters. MBs (~3 μm diameter) were repeatedly compressed for deformations up to ~50% to obtain a full bubble response. Addition of a coating increased the initial deformation stiffness related to shell bending ~2-fold for streptavidin and ∼3-fold for Q-dots. The presence of a polyethylene glycol (PEG) linker in between the lipid and functional coating, led to enhanced stiffening at high deformations. The plasticity index has been determined and only those MBs that included the PEG linker showed a force dependent short time-scale (<~1s) plasticity. This study demonstrates modulation of the mechanical response of biocompatible MBs through the addition of functional coatings necessary for rationale design of therapeutic lipid MBs for targeted drug delivery.

On-chip determination of C-reactive protein using magnetic particles in continuous flow

We demonstrate the application of a multilaminar flow platform, in which functionalized magnetic particles are deflected through alternating laminar flow streams of reagents and washing solutions via an external magnet, for the rapid detection of the inflammatory biomarker, C-reactive protein (CRP). The two-step sandwich immunoassay was accomplished in less than 60 s, a vast improvement on the 80-300 min time frame required for enzyme-linked immunosorbent assays (ELISA) and the 50 min necessary for off-chip magnetic particle-based assays. The combination of continuous flow and a stationary magnet enables a degree of autonomy in the system, while a detection limit of 0.87 μg mL(-1) makes it suitable for the determination of CRP concentrations in clinical diagnostics. Its applicability was further proven by assaying real human serum samples and comparing those results to values obtained using standard ELISA tests.

Simultaneous trapping of magnetic and diamagnetic particle plugs for separations and bioassays

We demonstrate the application of magnetic forces for the simultaneous trapping of two types of particles, magnetic and diamagnetic, via a single set of magnets. Furthermore, we show how this simple setup can be employed for performing assays with a negative control, or for achieving multiple simultaneous analyses.

Diamagnetic repulsion of particles for multilaminar flow assays

We demonstrate diamagnetic repulsion forces for performing continuous multilaminar flow assays on particles based on their intrinsic properties and with a simple setup. The platform could be applied to sandwich assays on polystyrene particles, and to cell-based assays via their suspension in biologically benign magnetic media.

Research spotlight: microbubbles for therapeutic delivery.

Systemic injection of chemotherapy agents for treating cancer can cause severe side effects for the patient, as well as being a relatively inefficient use of expensive and highly toxic drugs. The area of targeted drug delivery in which drugs are delivered utilizing a specialized carrier directly to the cancerous tumor via immuno-recognition has gained much interest in recent years. Such an approach reduces the side effects of systemic injection and also provides a localized, high-concentration treatment directly to the cancer. Our group at the University of Leeds (Leeds, UK) is developing therapeutic microbubbles that double as both agents for contrast-enhanced ultrasound imaging and drug-delivery vehicles that are targeted to specific cancer cell receptors. Ultimately, a large amplitude sound wave will be used to destroy the bubbles and trigger release of the drug at the targeted tumor. Theranostic microbubbles are a simple and versatile drug-delivery technique that could potentially improve cancer treatment, both in terms of patient experience and overall drug efficiency. Importantly, they offer new ways of delivering hydrophobic drugs, which have traditionally been difficult to deliver efficiently.

Single molecule protein biophysics using chemically modified nanopores

The kinetics of protein folding and binding are not only scientifically relevant to understanding the complex molecular machine-like functionality of proteins inside of cells but can also help elucidate disease pathways and lead to better therapeutic agents. Using nanopores to investigate these kinetics holds great potential for such proteomic studies in which the structure and function of proteins can be rapidly screened. In this study, we achieve part of this goal by detecting the folded and unfolded states of BSA. Furthermore, we also show that protein sensing can be performed on more biologically significant protein domains such as PDZ2. To achieve this goal, pore fabrication methods and chemical surface modifications were investigated and optimized for efficient protein sensing.

Evaluation of Lipid-Stabilised Tripropionin Nanodroplets as a Delivery Route for Combretastatin A4

Lipid-based nanoemulsions are a cheap and elegant route for improving the delivery of hydrophobic drugs. Easy and quick to prepare, nanoemulsions have promise for the delivery of different therapeutic agents. Although multiple studies have investigated the effects of the oil and preparation conditions on the size of the nanoemulsion nanodroplets for food applications, analogous studies for nanoemulsions for therapeutic applications are limited. Here we present a study on the production of lipid-stabilised oil nanodroplets (LONDs) towards medical applications. A number of biocompatible oils were used to form LONDs with phospholipid coatings, and among these, squalane and tripropionin were chosen as model oils for subsequent studies. LONDs were formed by high pressure homogenisation, and their size was found to decrease with increasing production pressure. When produced at 175MPa, all LONDs samples exhibited sizes between 100 and 300nm, with polydispersity index PI between 0.1 and 0.3. The LONDs were stable for over six weeks, at 4°C, and also under physiological conditions, showing modest changes in size (<10%). The hydrophobic drug combretastatin A4 (CA4) was encapsulated in tripropionin LONDs with an efficiency of approximately 76%, achieving drug concentration of approximately 1.3mg/ml. SVR mouse endothelial cells treated with CA4 tripropionin LONDs showed the microtubule disruption, characteristic of drug uptake for all tested doses, which suggests successful release of the CA4 from the LONDs.