Di-4-ANEPPDHQ, a fluorescent probe for the visualisation of membrane microdomains in living Arabidopsis thaliana cells

Abstract

Cholesterol-enriched microdomains, also called lipid rafts, are nanoscale membrane structures with a high degree of structural order. Since these microdomains play important roles in dynamic cytological events, such as cell signalling and membrane trafficking, the detection and tracking of microdomain behaviours are crucial to studies on modern membrane physiology. Currently, observation of microdomains is mostly based on the detection of specific raft-resident constituents using artificial cross-link fluorescent probes. However, only a few microdomain-specific fluorescent dyes are available for plant cell biology studies. In this study, the photophysical properties of di-4-ANEPPDHQ were analysed. The use of confocal laser scanning microscope (CLSM)-based methods in the visualisation of microdomains in living cells of Arabidopsis thaliana was assessed. The results confirmed that the generalised polarisation (GP) method can be used to quantitatively visualise the membrane orders in live plant cells. This dye was found to have low cytotoxicity in plant root epidermal cells and root hairs. These findings suggest that di-4-ANEPPDHQ is an appropriate tool for the visualisation of microdomains in living plant cells.

Keywords: Arabidopsis; Di-4-ANEPPDHQ; Lipid order; Living cells; Microdomain; Quantitative imaging.