title: Fixation and Permeabilization Approaches for Scanning Electrochemical Microscopy of Living Cells.
abstract: Scanning electrochemical microscopy (SECM) has been widely used for the electrochemical imaging of dynamic topographical and metabolic changes in alive adherent mammalian cells. However, extracting intracellular information by SECM is challenging, since it requires redox species to travel in and out the lipid cell membrane. Herein, we present cell fixation and permeabilization approaches as an alternative tool for visualizing cell properties by SECM. With this aim, adherent cells were analyzed in the SECM feedback mode in three different conditions: (i) alive; (ii) fixed, and (iii) fixed and permeabilized. The fixation was carried out with formaldehyde and does not damage lipid membranes. Therefore, this strategy can be used for the SECM investigation of cell topography or the passive transport of the redox mediator into the cells. Additional permeabilization of the cell membrane after fixation enables the analysis of the intracellular content through the coupling of SECM with immunoassay strategies for the detection of specific biomarkers. The latter was successfully applied as an easy and fast screening approach to detect the expression of the melanoma-associated marker tyrosinase in adherent melanoma cell lines corresponding to different cancer progression stages using the SECM substrate generation-tip collection mode. The present approach is simple, fast, and reliable and can open new ways to analyze cell cultures with electrochemically based scanning probe techniques.
authors.affiliation: Laboratoire d'Electrochimie Physique et Analytique, EPFL Valais Wallis, École Polytechnique Fédérale de Lausanne , CH-1951 Sion, Switzerland.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Bondarenko
authors.affiliation: Laboratoire d'Electrochimie Physique et Analytique, EPFL Valais Wallis, École Polytechnique Fédérale de Lausanne , CH-1951 Sion, Switzerland.
authors.forename: Tzu-En
authors.initials: TE
authors.lastname: Lin
authors.affiliation: Laboratory of the Physics of Living Matter, École Polytechnique Fédérale de Lausanne , CH-1015 Lausanne, Switzerland.
authors.forename: Petar
authors.initials: P
authors.lastname: Stupar
authors.affiliation: Laboratoire d'Electrochimie Physique et Analytique, EPFL Valais Wallis, École Polytechnique Fédérale de Lausanne , CH-1951 Sion, Switzerland.
authors.forename: Andreas
authors.initials: A
authors.lastname: Lesch
authors.affiliation: Laboratoire d'Electrochimie Physique et Analytique, EPFL Valais Wallis, École Polytechnique Fédérale de Lausanne , CH-1951 Sion, Switzerland.
authors.forename: Fernando
authors.initials: F
authors.lastname: Cortés-Salazar
authors.affiliation: Laboratoire d'Electrochimie Physique et Analytique, EPFL Valais Wallis, École Polytechnique Fédérale de Lausanne , CH-1951 Sion, Switzerland.
authors.forename: Hubert H
authors.initials: HH
authors.lastname: Girault
authors.affiliation: Laboratory of Physical Chemistry of Polymers and Membranes, École Polytechnique Fédérale de Lausanne , CH-1015 Lausanne, Switzerland.
authors.forename: Horst
authors.initials: H
authors.lastname: Pick
mesh_terms: D014408:Biomarkers, Tumor / Q000032:analysis* / Q000378:metabolism; D002470:Cell Survival; D005557:Formaldehyde / Q000737:chemistry; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D007118:Immunoassay; D064347:Microscopy, Electrochemical, Scanning; D014442:Monophenol Monooxygenase / Q000032:analysis* / Q000378:metabolism; D010084:Oxidation-Reduction; D014407:Tumor Cells, Cultured
chemical_list: D014408:Biomarkers, Tumor; D005557:Formaldehyde; D014442:Monophenol Monooxygenase