Magnetic tweezers are powerful tools to manipulate and study the mechanical properties of biological molecules and living cells. In this paper we present a novel, bona fide electromagnetic tweezer (EMT) setup that allows independent control of the force and torque applied via micrometer-sized magnetic beads to a molecule under study. We implemented this EMT by combining a single solenoid that generates force (f-EMT) with a set of four solenoids arranged into a symmetric quadrupole to generate torque (τ-EMT). To demonstrate the capability of the tweezers, we attached optically asymmetric Janus beads to single, tethered DNA molecules. We show that tension in the piconewton force range can be applied to single DNA molecules and the molecule can simultaneously be twisted with torques in the piconewton-nanometer range. Furthermore, the EMT allows the two components to be independently controlled. At various force levels applied to the Janus bead, the trap torsional stiffness can be continuously changed simply by varying the current magnitude applied to the τ-EMT. The flexible and independent control of force and torque by the EMT makes it an ideal tool for a range of measurements where tensional and torsional properties need to be studied simultaneously on a molecular or cellular level.
The ability of two, scattering gold nanoparticles (GNPs) to plasmonically couple in a manner that is dependent on the interparticle separation has been exploited to measure nanometer-level displacements. However, despite broad applicability to monitoring biophysical dynamics, the long time scales (<5 Hz) with which plasmonic coupling are typically measured are not suitable for many dynamic molecular processes, generally occurring over several milliseconds. Here, we introduce a new technique intended to overcome this technical limitation: ratiometric analysis using monochromatic, evanescent darkfield illumination (RAMEDI). As a proof-of-principle, we monitored dynamic, plasmonic coupling arising from the binding of single biotin- and neutravidin-GNPs with a temporal resolution of 38 ms. We also show that the observable bandwidth is extendable to faster time scales by demonstrating that RAMEDI is capable of achieving a signal-to-noise ratio greater than 20 from individual GNPs observed with 200 Hz bandwidth.
Atomic force microscopy (AFM) is a powerful tool for imaging and chemical characterization of bio-samples at molecular resolution in physiologicallyrelevant environments. However, the localized tip-sample interactions limit high-resolution images to the topmost layer of surfaces. Consequently, characterizing the three-dimensional (3-D) inner structures of molecules has been a challenge. Here, we demonstrate three-dimensional (3-D) localization of chemical groups within a single protein complex using AFM. We employ short DNA sequences to label specific chemical groups inside a protein complex. T-shaped cantilevers functionalized with complementary probe DNAs allow locating each label with sequence specificity and sub-nanometer resolution. We also measure pairwise distances between labels and reconstruct the 3-D loci of the target groups using simple geometric calculations. Experiments with the biotin-streptavidin complex showed that the 3-D loci of carboxylic acids of biotins are within 2-Angstroms of their respective 3-D loci in the corresponding crystal structure, suggesting AFM may complement existing structural biological techniques in solving structures that are difficult to study due to their size and complexity. This technique maybe finds applications in studying structure of DNA or RNA binding proteins.
Luminescence is ubiquitous in biology research and medicine. Conceptually simple, the detection of luminescence nonetheless faces technical challenges because relevant signals can exhibit exceptionally low radiant power densities. Although low light detection is well-established in centralized laboratory settings, the cost, size, and environmental requirements of high-performance benchtop luminometers are not compatible with geographically-distributed global health studies or resource-constrained settings. Here we present the design and application of a ~$700 US handheld, battery-powered luminometer with performance on par with high-end benchtop instruments. By pairing robust and inexpensive Silicon Photomultiplier (SiPM) sensors with a low-profile shutter system, our design compensates for sensor non-idealities and thermal drift, achieving a limit of detection of 1.6E-19 moles of firefly luciferase, or approximately 1fW of radiant optical power. Using these devices, we performed split luciferase serology studies to monitor sars-cov-2 antibodies in a cohort in the United States, as well as a field study in Bangladesh.
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