Nanoscopy

Molecules can be readily designed by chemists, and a multitude of time-tested techniques are available for examining whether or not the correct molecule was made. However, when we begin to transition away from small molecules, to macromolecules, nanomaterials, and beyond, we have a larger characterization problem. Specifically, we require combinations of methods that can yield chemical information, and overall morphological data. The nanoscale interface – where molecules become materials – is a fascinating length and time scale to work at, simply because nanomaterials are too large for standard chemistry techniques, too small for light microscopy, and move too quickly to capture. We are broadly interested in the characterization challenges presented by complex, dynamic nanoscale materials, and seek to develop new strategies, and combine methods to yield information.

In Situ TEM of Soft Matter

The formation and function of many of the most essential nanoscale structures requires them to be solvated in specific liquids, from the ubiquitous biological supramolecular assemblies that are essential to life, to modern synthetic nanomaterials. For such systems, conventional dry-state Transmission Electron Microscopy (TEM) is limited in its ability to probe these solution phase structures and will never give us the ability to observe them dynamically, in motion. Dynamics are at the heart of their functionality, including nucleation and growth processes, and complex phase changes in response to external stimuli (e.g. heat, pH, enzyme). We use a liquid cell inside a TEM, which seals picoliter volumes of liquid in a compartment which withstands the vacuum of the microscope, to directly observe and video processes in real time with nanometer resolution. This data often provides us with a view into a length domain previously inaccessible.

In particular, we are interested in soft matter-based systems, including but not limited to proteins, polymers, inorganic/organic hybrid nanoparticles, metal-organic frameworks, and biological materials. To make LCTEM a viable characterization method for most laboratories, we are also interested in developing LCTEM technology to allow for robust, controlled, and reproducible LCTEM experiments and to establish chemical gradients and the ability to mix materials for visualization of reactions and reactivity.

Observing the Growth of Metal-Organic Frameworks by In-situ Liquid Cell TEM
Joseph P. Patterson, Patricia Abellan, Michael S. Denny, Jr., Chiwoo Park, Nigel D. Browning, Seth M. Cohen, James E. Evans, and Nathan C. Gianneschi
J. Am. Chem. Soc., 2015, 137 (23), 7322–7328, DOI: 10.1021/jacs.5b00817

Colloidal Covalent Organic Frameworks
Brian Smith,^† Lucas Parent,^† Anna Overholts, Peter Beaucage, Ryan Bisbey, Anton Chavez, Nicky Hwang, Chiwoo Park, Austin Evans, Nathan C. Gianneschi,* William Dichtel*
ACS Central Science, 2017, 3 (1), 58–65, DOI: 10.1021/acscentsci.6b00331

Picoliter Drop-on-Demand Dispensing for Multiplex Liquid Cell TEM
Joseph P. Patterson,^† Lucas R. Parent,^† Joshua Cantlon, Holger Eickhoff, Guido Bared, James E. Evans, and Nathan C. Gianneschi*
Microscopy and Microanalysis, 2016, 22 (3), 507–514, DOI: 10.1017/S1431927616000659

Dynamics of Soft Nanomaterials Captured by Transmission Electron Microscopy in Liquid Water
Maria T. Proetto, Anthony M. Rush, Miao-Ping Chien, Patricia Abellan Baeza, Joe P. Patterson, Matthew P. Thompson, Norman H. Olson, Curtis E. Moore, Arnold L. Rheingold, Christopher Andolina, Jill Millstone, Stephen B. Howell, Nigel D. Browning, James E. Evans, and Nathan C. Gianneschi
J. Am. Chem. Soc., 2014, 136 (4), 1162-1165, DOI: 10.1021/ja408513m

Multi-Scale and Multimodal Microscopies

The design of new nanomedicines depends on an intimate understanding of the nanostructure itself and how it interacts with complex biological systems. We design labeled nanoprobes and employ super-resolution imaging techniques, as well multi-modal approaches, to fully understand these interactions:

Nanoscale Secondary Ion Mass Spectrometry (NanoSIMS)
Structured Illumination Microscopy (SIM)
Stochastic Optical Reconstruction Microscopy (STORM)
Biological-TEM

Correlated optical and isotopic nanoscopy (COIN) has been recently introduced for the study of biological samples such as cells and tissues. With this idea in mind we used NanoSIMS (Nanoscale Secondary Ion Mass Spectrometry) and SIM (Structureal Illumination Microscopy) to study isotopically and fluorescently labeled nanomaterials in biological systems such as cells and tissues. This method allowed us not only to track single nanoparticles and study their composition by NanoSIMS, but at the same time visualize their subcellular localization at different stages of their uptake and metabolization process by identifying different cellular organelles using fluorescent labels for SIM. Super-resolution fluorescence microscopy techniques such as STORM (Stochastic Optical Reconstruction Microscopy) enable the examination of submicrometer structures by overcoming the diffraction of light limit. Transmission Electron Microscopy (TEM) is widely applied to study ultrastructure of the cells and tissues with nanometric resolution. With this propose thin slices of generally plastic-embedded stained samples are placed under an electron beam and an image is obtained from the specific contrast given by different organelles in the cell.

Cellular Delivery of Nanoparticles Revealed with Combined Optical and Isotopic Nanoscopy
Maria T. Proetto, Christopher R. Anderton, Dehong Hu, Craig J. Szymanski, Zihua Zhu, Joseph P. Patterson, Jacquelin Kammeyer, Lizanne G. Nilewski, Anthony M. Rush, Nia C. Bell, James E. Evans, Galya Orr, Stephen B. Howell, and Nathan C. Gianneschi*
ACS Nano, 2016, 10 (4), 4046–4054, DOI: 10.1021/acsnano.5b06477

Enzyme-Directed Assembly of Nanoparticles in Tumors Monitored by In Vivo Whole Animal and Ex Vivo Super Resolution Fluorescence Imaging
Miao-Ping Chien, Andrea S. Carlini, Dehong Hu, Christopher V. Barback, Anthony M. Rush, David J. Hall, Galya Orr, and Nathan C. Gianneschi*
J. Am. Chem. Soc., 2013, 135 (50), 18710-18713, DOI: 10.1021/ja408182p

Mimicking Melanosomes: Polydopamine Nanoparticles as Artificial Microparasols
Yuran Huang, Yiwen Li , Ziying Hu, Xiujun Yue, Maria T. Proetto, Ying Jones, and Nathan C. Gianneschi*
ACS Cent. Sci., 2017, 3(6), 564–569, DOI: 10.1021/acscentsci.6b00230

Cryo-EM of Biomaterials

Cryogenic electron microscopy (cryo-EM) is a characterization method that has revolutionized the field of structural biology. In cryo-EM, a thin liquid film of sample is vitrified in liquid ethane, then stored in liquid nitrogen and imaged at <-175°C. In doing so, the sample’s native conformation and structure is maintained, and can be imaged with sub-Ångstrom resolution. In addition, no staining agent is required, even for extremely low-contrast, low-scattering materials. This is especially important for non-crystalline materials, which otherwise undergo morphology changes upon drying, or upon staining with heavy metal salts. In the Gianneschi Lab, we utilize cryo-EM for imaging natural and synthetic soft nanomaterials.

Sea spray aerosol structure and composition using cryogenic transmission electron microscopy
Joseph Patterson*, Douglas Collins, Jennifer Michaud, Jessica Axson, Camille Sultana,Trever Moser, Abigail Dommer, Jack Conner, Vicki Grassian, M. Stokes, Grant Deane, James Evans, Michael Burkart, Kimberly Prather, Nathan C. Gianneschi
ACS Central Science, 2016, 2 (1), 40–47, DOI: 10.1021/acscentsci.5b00344

Phase Diagrams of Polynorbornene Amphiphilic Block Copolymers in Solution
Sarah A. Barnhill, Nia C. Bell, Joseph P. Patterson, Daniel P. Olds, and Nathan C. Gianneschi* Macromolecules, 2015, 48 (4), 1152–1161, DOI: 10.1021/ma502163j

Analytical (S)TEM

(Scanning) Transmission Electron Microscopes are not only able to image nanomaterials with sub-Ångstrom resolution, but when equipped with spectrometers and energy filters, are able to probe and even quantify their chemical composition with location specificity, a characterization technique called Analytical (S)TEM. We use two varieties of analytical STEM, both of which acquire signal from atom ionization events caused by the incident e- beam; 1) collecting characteristic emitted X-rays (energy dispersive x-ray spectroscopy, EDS) or 2) measuring the energy loss of the incident e- beam (electron energy loss spectroscopy, EELS). We then use collected spectroscopic signal to create elemental maps of nanostructures to pair with the conventional (S)TEM images of their morphology. For samples that can be characterized in high-vacuum (dry-sate or cryo-vitrified), this is among the most powerful nanoscale characterization techniques, which we use to verify the presence of loaded metals in polymeric nanoparticles, and to map and quantify the precise elemental composition of metal-organic frameworks (MOFs).

Pore Breathing of Metal-Organic Frameworks by Environmental Transmission Electron Microscopy
Lucas R. Parent,^† Huy Pham,^† Joseph P. Patterson, Michael S Denny, Jr., Seth M. Cohen,* Nathan C. Gianneschi,* and Francesco Paesani*
J. Am. Chem. Soc., 2017, 139 (40), 13973–13976, DOI: 10.1021/jacs.7b06585

Tunable, Metal-Loaded Polydopamine Nanoparticles Analyzed by Magnetometry
Zhao Wang,^† Yijun Xie,^† Yiwen Li, Yuran Huang, Lucas R. Parent, Treffly Ditri, Nanzhi Zang, Jeffrey D. Rinehart,* and Nathan C. Gianneschi*
Chemistry of Materials, 2017, 29 (19), 8195–8201, DOI: 10.1021/acs.chemmater.7b02262