info:eu-repo/semantics/article
Quantitative expansion microscopy for the characterization of the spectrin periodic skeleton of axons using fluorescence microscopy
Fecha
2020-02Registro en:
Martínez, Gaby F.; Gazal, Nahir Guadalupe; Quassollo Infanzon, Gonzalo Emiliano; Szalai, Alan Marcelo; Del Cid Pellitero, Esther; et al.; Quantitative expansion microscopy for the characterization of the spectrin periodic skeleton of axons using fluorescence microscopy; Nature Publishing Group; Scientific Reports; 10; 1; 2-2020; 1-11
2045-2322
2045-2322
CONICET Digital
CONICET
Autor
Martínez, Gaby F.
Gazal, Nahir Guadalupe
Quassollo Infanzon, Gonzalo Emiliano
Szalai, Alan Marcelo
Del Cid Pellitero, Esther
Durcan, Thomas M.
Fon, Edward A.
Bisbal, Mariano
Stefani, Fernando Daniel
Unsain, Nicolas
Resumen
Fluorescent nanoscopy approaches have been used to characterize the periodic organization of actin,spectrin and associated proteins in neuronal axons and dendrites. This membrane-associated periodicskeleton (MPS) is conserved across animals, suggesting it is a fundamental component of neuronalextensions. The nanoscale architecture of the arrangement (190 nm) is below the resolution limitof conventional fluorescent microscopy. Fluorescent nanoscopy, on the other hand, requires costlyequipment and special analysis routines, which remain inaccessible to most research groups. Thisreport aims to resolve this issue by using protein-retention expansion microscopy (pro-ExM) to revealthe MPS of axons. ExM uses reagents and equipment that are readily accessible in most neurobiologylaboratories. We first explore means to accurately estimate the expansion factors of protein structureswithin cells. We then describe the protocol that produces an expanded specimen that can be examinedwith any fluorescent microscopy allowing quantitative nanoscale characterization of the MPS. Wevalidate ExM results by direct comparison to stimulated emission depletion (STED) nanoscopy. Weconclude that ExM facilitates three-dimensional, multicolor and quantitative characterization of theMPS using accessible reagents and conventional fluorescent microscopes.