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MicroscopyInnovations


Previously, micro-scale observations of benthic marine specimens could only be performed on samples in the lab under a bench-top microscope. T is signifi cantly limited the types of processes that could be observed. Ocean habitats are complex settings, and many interactions occur between diff erent species; additionally the organisms interact with a dynamic physical environment that has continuous fl uctuations in parameters such as temperature, fl uid motion, and pH. T is environment is thus exceedingly diffi cult to fully simulate in the lab.


Scanning Thermo-Ionic Microscopy (STIM)


University of Washington and Shenzhen Institutes of Advanced Technology


Developers: Jiangyu Li, Ahmadreza Eshghinejad, and Ehsan Nasr Esfahani


Scanning thermo-ionic microscopy (STIM) probes local electrochemical activities via Vegard strain induced by thermal stress. Vegard strain is a lattice volume change resulting from a change in the concentration of an ionic species.


T is strain can be produced by local temperature fl uctuations induced either by resistive heating or photo-thermal heating, resulting in thermal expansion and a stress that drives ionic oscillation. T e Vegard strain resulting from this ionic oscillation can then be detected at appropriate harmonics, enabling sensitive detection with high spatial resolution. T is is the essence of STIM. T e dynamics of ionic motion can be captured from point-wise spectroscopy studies, while spatial inhomogeneity can be revealed by STIM mapping.


Implementation of this new method has been demonstrated


on modifi ed commercial atomic force microscopes (AFMs). Detection of the harmonic responses of cantilever defl ection, usually very small in magnitude, were accomplished by a lock-in amplifi er around the cantilever-sample contact resonance frequency, which enhances the signal-to-noise ratio by orders of magnitude. To avoid topography cross-talk during STIM scanning, a dual amplitude resonance tracking (DART) technique was used to track the contact resonance. Conventional electrochemical characterization techniques


are very diffi cult to scale down because they are mostly based on current measurement, requiring the detection of small currents on the order of picoamperes at the nanoscale. An earlier scanning probe technique of electrochemical strain microscopy (ESM) off ers high sensitivity and spatial resolution in exciting Vegard strain electrically. However, it is oſt en diffi cult to distinguish Vegard strain from other electromechanical mechanisms such as the piezoelectric eff ect, electrostatic interactions, and capacitive forces. STIM overcomes this diffi culty by using Vegard strain excited by stress instead of an electric fi eld, and it still provides high sensitivity and spatial resolution similar to ESM. Since excitation is not electrical in nature—eliminating the interference from other electrochemical mechanisms— in operando testing becomes possible. T e STIM method can be applied to study a wide range of electrochemical materials including electrode materials for


2017 September • www.microscopy-today.com


battery systems, fuel cells and fl ow batteries, electrocatalysts for effi cient electrosynthesis of liquid fuels, and photoelectrochemical materials that can directly convert solar energy to fuels.


Multimodal Imaging with a Coherence Switching Laser


Yale University


Developers: Sebastian Knitter, Changgeng Liu, Brandon Redding, Mustafa K. Khokha, Michael A. Choma, and Hui Cao


Unlike the traditional laser sources with a fixed degree of high spatial coherence, degenerate vertical external-cavity surface-emitting lasers (VECSELs) can have a tunable degree of spatial coherence, which is a new degree of freedom in microscopy. This new laser source can switch between emission of two different spatial coherence states: high spatial coherence and low spatial coherence.


These two coherence states yield different kinds of imaging contrast in light microscopy, and the ability to perform coherence switching enables multimodal microscopic imaging. Low-spatial-coherence illumination enables speckle-free imaging of structures in specimens. High-spatial- coherence illumination enables dynamic speckle imaging of moving objects and blood flow in specimens. In particular, speckle fluctuation analysis yields metrics that are related to blood flow speed. This contrast is endogenous in nature and does not require the use of external contrast agents. A degenerate laser has a self-imaging laser cavity; one end of the resonator cavity is imaged onto the other end of the cavity. By placing a wide-area gain element at one end of the cavity, the resonator supports a large number (~1000) of non-overlapping, mutually incoherent modes. For our laser, a wide-area electrically- pumped semiconductor VECSEL was used as the gain element. A large number of transverse lasing modes combine to generate low- spatial-coherence (speckle-free) emission. T e use of a spatial fi lter within the self-imaging cavity can eff ectively phase-lock the large number of modes into a single or a few spatial modes while maintaining emission power and direction. A degenerate laser with a VECSEL gain element also has the advantages of relatively low cost and uncomplicated thermal management. Additionally, degenerate lasers are much brighter than traditional light sources used in microscopy (for example, thermal sources and LEDs). T e ability to perform multimodal imaging was demonstrated in a frog embryo (a tadpole, an important animal model in congenital heart disease research). Dynamic structural imaging of the embryo heart was performed using speckle-free, low-spatial-coherence illumination, whereas functional speckle fl ow imaging was accomplished with high-spatial-coherence illumination. T is imaging demonstrated heart wall and blood fl ow dynamics over the cardiac cycle (that is, the heart fi lling with blood and the heart ejecting blood to the body).


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