Results for “Total Internal Reflection Microscopy”

DOE OSTI2025

Cryosectioning-enhanced super-resolution microscopy for single-protein imaging across cells and tissues

DNA-points accumulation for imaging in nanoscale topography (DNA-PAINT) enables nanoscale imaging with virtually unlimited multiplexing and molecular counting. Here, we address challenges, such as variable imaging performance and target accessibility, that can limit its broader applicability. Specifically, we enhance its capacity for robust single-protein imaging and molecular counting by optimizing the integration of total internal reflection fluorescence microscopy with physical sectioning, in particular, Tokuyasu cryosectioning. Our method, tomographic and kinetically enhanced DNA-PAINT (tkPAINT), achieves 3 nm localization precision across diverse samples, enhanced imager binding, and improved cellular integrity. tkPAINT can facilitate molecular counting with DNA-PAINT inside the nucleus, as demonstrated through its quantification of the in situ abundance of RNA Polymerase II in both HeLa cells as well as mouse tissues. Anticipating that tkPAINT could become a versatile tool for the exploration of biomolecular organization and interactions across cells and tissues, we also demonstrate its capacity to support multiplexing, multimodal targeting of proteins and nucleic acids, and three-dimensional (3D) imaging.

Science & Technology - Other Topics↗

DOE OSTI2025

An Internal Digital Image Correlation Technique for High-Strain Rate Dynamic Experiments

Full-field, quantitative visualization techniques, such as digital image correlation (DIC), have unlocked vast opportunities for experimental mechanics. However, DIC has traditionally been a surface measurement technique, and has not been extended to perform measurements on the interior of specimens for dynamic, full-scale laboratory experiments. This limitation restricts the scope of physics which can be investigated through DIC measurements, especially in the context of heterogeneous materials. The focus of this study is to develop a method for performing internal DIC measurements in dynamic experiments. The aim is to demonstrate its feasibility and accuracy across a range of stresses (up to 650 MPa), strain rates (10 3 - 10 6 s -1 ), and high-strain rate loading conditions (e.g., ramped and shock wave loading). Internal DIC is developed based on the concept of applying a speckle pattern at an inner-plane of a transparent specimen. The high-speed imaging configuration is coupled to the traditional dynamic experimental setups, and is focused on the internal speckle pattern. During the experiment, while the sample deforms dynamically, in-plane, two-dimensional deformations are measured via correlation of the internal speckle pattern. In this study, the viability and accuracy of the internal DIC technique is demonstrated for split-Hopkinson (Kolsky) pressure bar (SHPB) and plate impact experiments. The internal DIC experimental technique is successfully demonstrated in both the SHPB and plate impact experiments. In the SHPB setting, the accuracy of the technique is excellent throughout the deformation regime, with measurement noise of approximately 0.2% strain. In the case of plate impact experiments, the technique performs well, with error and measurement noise of 1% strain. The internal DIC technique has been developed and demonstrated to work well for full-scale dynamic high-strain rate and shock laboratory experiments, and the accuracy is quantified. Here, the technique can aid in investigating the physics and mechanics of the dynamic behavior of materials, including local deformation fields around dynamically loaded material heterogeneities.

36 MATERIALS SCIENCE↗