Single-molecule FRET combined with electrokinetic trapping reveals real-time enzyme kinetics of individual F-ATP synthases.

Single-molecule Förster resonance energy transfer (smFRET) is a key technique to observe conformational changes in molecular motors and to access the details of single-molecule static and dynamic disorder during catalytic processes. However, studying freely diffusing molecules in solution is limited to a few tens of milliseconds, while surface attachment often bears the risk to restrict their natural motion. In this paper we combine smFRET and electrokinetic trapping (ABEL trap) to non-invasively hold single FF-ATP synthases for up to 3 s within the detection volume, thereby extending the observation time by a factor of 10 as compared to Brownian diffusion without surface attachment.

Single-molecule FRET dynamics of molecular motors in an ABEL trap.

Single-molecule Förster resonance energy transfer (smFRET) of molecular motors provides transformative insights into their dynamics and conformational changes both at high temporal and spatial resolution simultaneously. However, a key challenge of such FRET investigations is to observe a molecule in action for long enough without restricting its natural function. The Anti-Brownian ELectrokinetic Trap (ABEL trap) sets out to combine smFRET with molecular confinement to enable observation times of up to several seconds while removing any requirement of tethered surface attachment of the molecule in question.

Probing Ligand-Receptor Interaction in Living Cells Using Force Measurements With Optical Tweezers.

This work probes the binding kinetics of COOH-terminus of (c-CPE) and claudin expressing MCF-7 cells using force spectroscopy with optical tweezers. c-CPE is of high biomedical interest due to its ability to specifically bind to claudin with high affinity as well as reversibly disrupt tight junctions whilst maintaining cell viability. We observed single-step rupture events between silica particles functionalized with c-CPE and MCF-7 cells.

Optically trapped bacteria pairs reveal discrete motile response to control aggregation upon cell-cell approach.

Aggregation of bacteria plays a key role in the formation of many biofilms. The critical first step is cell-cell approach, and yet the ability of bacteria to control the likelihood of aggregation during this primary phase is unknown. Here, we use optical tweezers to measure the force between isolated Bacillus subtilis cells during approach.

Measuring nanoparticle flow with the image structure function.

We present a technique to measure the velocity and flow profiles of a nanofluid in a microfluidic channel. Importantly, we extract the flow velocity from a series of standard brightfield images without employing particle tracking or laser-enhanced methods. Our analysis retrieves the flow information from the image structure function of sub-diffraction limited nanoparticles in suspension.

Holographic aberration correction: optimising the stiffness of an optical trap deep in the sample.

We investigate the effects of 1(st) order spherical aberration and defocus upon the stiffness of an optical trap tens of μm into the sample. We control both these aberrations with a spatial light modulator. The key to maintain optimum trap stiffness over a range of depths is a specific non-trivial combination of defocus and axial objective position.

Optical vortex trap for resonant confinement of metal nanoparticles.

The confinement and controlled movement of metal nanoparticles and nanorods is an emergent area within optical micromanipulation. In this letter we experimentally realise a novel trapping geometry near the plasmon resonance using an annular light field possessing a helical phasefront that confines the nanoparticle to the vortex core (dark) region. We interpret our data with a theoretical framework based upon the Maxwell stress tensor formulation to elucidate the total forces upon nanometric particles near the particle plasmon resonance.