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Ultramicroscopy. 2016 Jan;160:168-181. doi: 10.1016/j.ultramic.2015.10.014. Epub 2015 Oct 19.

Synchronizing atomic force microscopy force mode and fluorescence microscopy in real time for immune cell stimulation and activation studies.

Author information

1
Aix Marseille Université, LAI UM 61, Marseille F-13288, France; Inserm, UMR_S 1067, Marseille F-13288, France; CNRS, UMR 7333, Marseille F-13288, France.
2
Aix Marseille Université, LAI UM 61, Marseille F-13288, France; Inserm, UMR_S 1067, Marseille F-13288, France; CNRS, UMR 7333, Marseille F-13288, France; APHM, Hôpital de la Conception, Laboratoire d'Immunologie, Marseille F-13385, France.
3
Aix Marseille Université, LAI UM 61, Marseille F-13288, France; Inserm, UMR_S 1067, Marseille F-13288, France; CNRS, UMR 7333, Marseille F-13288, France. Electronic address: pierre-henri.puech@inserm.fr.

Abstract

A method is presented for combining atomic force microscopy (AFM) force mode and fluorescence microscopy in order to (a) mechanically stimulate immune cells while recording the subsequent activation under the form of calcium pulses, and (b) observe the mechanical response of a cell upon photoactivation of a small G protein, namely Rac. Using commercial set-ups and a robust signal coupling the fluorescence excitation light and the cantilever bending, the applied force and activation signals were very easily synchronized. This approach allows to control the entire mechanical history of a single cell up to its activation and response down to a few hundreds of milliseconds, and can be extended with very minimal adaptations to other cellular systems where mechanotransduction is studied, using either purely mechanical stimuli or via a surface bound specific ligand.

KEYWORDS:

Activation; Atomic force microscopy; Cell mechanics; Fluorescence microscopy; Immune cells; Signalling

PMID:
26521163
DOI:
10.1016/j.ultramic.2015.10.014
[Indexed for MEDLINE]
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