Protocol to measure centrosome cohesion deficits mediated by pathogenic LRRK2 in cultured cells using confocal microscopy

Abstract

Summary: The present protocol allows for quantification of inter-centrosome distances in G2 phase cells by confocal fluorescence microscopy to determine centrosome cohesion deficits. We describe transfection and immunofluorescence approaches followed by image acquisition and analysis of inter-centrosome distances. This protocol is for adherent A549 cells transiently overexpressing pathogenic LRRK2 and for immortalized murine embryonic fibroblasts endogenously expressing LRRK2 but is amenable to any other cultured cell type as well.For complete details on the use and execution of this protocol, please refer to Fdez et al.1 and Lara Ordóñez et al.2 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Keywords

• Cell Biology
• Cell Culture
• Cell-based Assays
• Microscopy