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1. Immunofluorescence Microscopy

   https://doi.org/10.1002/cpz1.842  

   Galati, Domenico F.; Asai, David J. (August 2023, Current Protocols)

   Abstract Visualizing fluorescence‐tagged molecules is a powerful strategy that can reveal the complex dynamics of the cell. One robust and broadly applicable method is immunofluorescence microscopy, in which a fluorescence‐labeled antibody binds the molecule of interest and then the location of the antibody is determined by fluorescence microscopy. The effective application of this technique includes several considerations, such as the nature of the antigen, specificity of the antibody, permeabilization and fixation of the specimen, and fluorescence imaging of the cell. Although each protocol will require fine‐tuning depending on the cell type, antibody, and antigen, there are steps common to nearly all applications. This article provides protocols for staining the cytoskeleton and organelles in two very different kinds of cells: flat, adherent fibroblasts and thick, free‐swimmingTetrahymenacells. Additional protocols enable visualization with widefield, laser scanning confocal, and eSRRF super‐resolution fluorescence microscopy. © 2023 Wiley Periodicals LLC. Basic Protocol 1: Immunofluorescence staining of adherent cells such as fibroblasts Basic Protocol 2: Immunofluorescence of suspension cells such asTetrahymena Basic Protocol 3: Visualizing samples with a widefield fluorescence microscope Alternate Protocol 1: Staining suspension cells adhered to poly‐l‐lysine‐coated coverslips Alternate Protocol 2: Visualizing samples with a laser scanning confocal microscope Alternate Protocol 3: Generating super‐resolution images with SRRF microscopy 

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2. Mechanical expansion microscopy

   https://doi.org/10.1016/bs.mcb.2020.04.013  

   Fan, Y.; Lim, Y.; Wyss, L. S.; Park, S.; Xu, C.; Fu, H.; Fei, J.; Hong, Y.; Wang, B. (January 2021, Methods in cell biology)

   Guichard, P.; Hamel, V. (Ed.)

   This chapter describes two mechanical expansion microscopy methods with accompanying step-by-step protocols. The first method, mechanically resolved expansion microscopy, uses non-uniform expansion of partially digested samples to provide the imaging contrast that resolves local mechanical properties. Examining bacterial cell wall with this method, we are able to distinguish bacterial species in mixed populations based on their distinct cell wall rigidity and detect cell wall damage caused by various physiological and chemical perturbations. The second method is mechanically locked expansion microscopy, in which we use a mechanically stable gel network to prevent the original polyacrylate network from shrinking in ionic buffers. This method allows us to use anti-photobleaching buffers in expansion microscopy, enabling detection of novel ultra-structures under the optical diffraction limit through super-resolution single molecule localization microscopy on bacterial cells and whole-mount immunofluorescence imaging in thick animal tissues. We also discuss potential applications and assess future directions. 

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3. scanning frequency comb microscopy: a new method in scanning probe microscopy

   https://doi.org/10.13140/RG.2.2.34585.72804  

   Hagmann, M. (August 2018, scanning frequency comb microscopy: a new method in scanning probe microscopy)

   Finer resolution with greater stability is possible using unique low-power (aW), low-noise (20 dB S/N), microwave harmonics generated within a nanoscale tip-sample junction for feedback control in place of the DC tunneling current. Please see the attached poster to be presented at the Microscopy & Microanalysis-2018 meeting in Baltimore Monday August 6th as Post-deadline poster PDP-18. Applications include true sub-nm resolution in the carrier profiling of semiconductors. This method is especially appropriate for resistive samples where the spreading resistance flattens plots of the tunneling current vs. tip-sample distance with a scanning tunneling microscope. 

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4. Scanning Ion Conductance Microscopy

   https://doi.org/10.1021/acs.chemrev.0c00962  

   Zhu, Cheng; Huang, Kaixiang; Siepser, Natasha P.; Baker, Lane A. (December 2020, Chemical Reviews)

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5. Indirect magnetic force microscopy

   https://doi.org/10.1039/c9na00193j  

   Sifford, Joshua; Walsh, Kevin J.; Tong, Sheng; Bao, Gang; Agarwal, Gunjan (June 2019, Nanoscale Advances)

   Magnetic force microscopy (MFM) is an atomic force microscopy (AFM)-based technique to map magnetic domains in a sample. MFM is widely used to characterize magnetic recording media, magnetic domain walls in materials, nanoparticles and more recently iron deposits in biological samples. However, conventional MFM requires multiple scans of the samples, suffers from various artifacts and is limited in its capability for multimodal imaging or imaging in a fluid environment. We propose a new modality, namely indirect magnetic force microscopy (ID-MFM), a technique that employs an ultrathin barrier between the probe and the sample. Using fluorescently conjugated superparamagnetic nanoparticles, we demonstrate how ID-MFM can be achieved using commercially available silicon nitride windows, MFM probes and AFM equipment. The MFM signals obtained using ID-MFM were comparable to those obtained using conventional MFM. Further, samples prepared for ID-MFM were compatible with multi-modal imaging via fluorescence and transmission electron microscopy. Thus ID-MFM can serve as a high-throughput, multi-modal microscopy technique which can be especially attractive for detecting magnetism in nanoparticles and biological samples. 

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