Angular Approach Scanning Ion Conductance Microscopy

Abstract

Scanning ion conductance microscopy (SICM) is a super-resolution live imaging technique that uses a glass nanopipette as an imaging probe to produce three-dimensional (3D) images of cell surface. SICM can be used to analyze cell morphology at nanoscale, follow membrane dynamics, precisely position an imaging nanopipette close to a structure of interest, and use it to obtain ion channel recordings or locally apply stimuli or drugs. Practical implementations of these SICM advantages, however, are often complicated due to the limitations of currently available SICM systems that inherited their design from other scanning probe microscopes in which the scan assembly is placed right above the specimen. Such arrangement makes the setting of optimal illumination necessary for phase contrast or the use of high magnification upright optics difficult. Here, we describe the designs that allow mounting SICM scan head on a standard patch-clamp micromanipulator and imaging the sample at an adjustable approach angle. This angle could be as shallow as the approach angle of a patch-clamp pipette between a water immersion objective and the specimen. Using this angular approach SICM, we obtained topographical images of cells grown on nontransparent nanoneedle arrays, of islets of Langerhans, and of hippocampal neurons under upright optical microscope. We also imaged previously inaccessible areas of cells such as the side surfaces of the hair cell stereocilia and the intercalated disks of isolated cardiac myocytes, and performed targeted patch-clamp recordings from the latter. Thus, our new, to our knowledge, angular approach SICM allows imaging of living cells on nontransparent substrates and a seamless integration with most patch-clamp setups on either inverted or upright microscopes, which would facilitate research in cell biophysics and physiology.

Hybrid scanning electrochemical cell microscopy-interference reflection microscopy (SECCM-IRM): tracking phase formation on surfaces in small volumes

“…28,29,31,95 Coverslips coated with graphene-based materials are also possible electrode candidates, as shown in the IRM imaging of the electrochemistry of graphene oxide. 33,95 Finally, reflectivity microscopy can also be operated at reflective and non-transparent electrodes 96 and the combination with a modified SECCM setup (e.g., with an angled nanopipette, as used in some scanning ion conductance microscopy studies), 97 could be explored. Among other applications, we shall also investigate direct electrodeposition (e.g.…”

Scanning Probe Microscopy for Nanoscale Electrochemical Imaging

“…Reproduced from Shevchuk, A.; Tokar, S.; Gopal, S.; Sanchez-Alonso, J. L.; Tarasov, A. I.; Velez-Ortega, A. C.; Chiappini, C.; Rorsman, P.; Stevens, M. M.; Gorelik, J.; Frolenkov, G. I.; Klenerman, D.; Korchev, Y. E. Biophys J. 2016 , 110 , 2252–2265 (ref ). Copyright 2016, with permission from Elsevier.…”

Advances in high-resolution microscopy for the study of intracellular interactions with biomaterials

“…Scanning probe microscopy techniques such as atomic force microscopy (AFM), scanning ion conductance microscopy (SICM), and scanning electron microscopy (SEM) have been highly effective at imaging extracellular biointerfaces at the nanoscale, demonstrating the impact of nanotopography on cell behavior [40][41][42]. However, the mechanisms that are involved in the cellular response to nanotopography, and the effect of surface chemistry on the cell, are less well studied.…”