Study of Diffusion of a polyelectrolyte in a multilayer film by Fluorescence recovery after photobleaching (FRAP) (CAT#: STEM-MT-0023-WXH)

Introduction

In recent years, the applications of polyelectrolyte multilayer films, built by alternating adsorption of anionic and cationic polyelectrolytes, have grown in various fields. Transport of drugs or active molecules coupled to the polyelectrolytes is of interest for biomedical applications, such as controlled release. In polyelectrolyte films having a thickness exponentially growing with the number of deposited layer pairs, at least one of the polyelectrolytes diffuses “in” and “out” of the films during their buildup.

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Principle Applications Procedure Materials Notes Related Products

Principle

Fluorescence recovery after photobleaching (FRAP) is a microscopy technique capable of quantifying the mobility of molecules within cells. By exploiting the phenomenon of photobleaching, fluorescent mole- cules within a region of interest can be selectively and irreversibly 'turned off'. It is capable of quantifying the two-dimensional lateral diffusion of a molecularly thin film containing fluorescently labeled probes, or to examine single cells.

Applications

• Characterization of the mobility of individual lipid molecules within a cell membrane.
• Analysis of molecule diffusion within the cell
• Study of protein interaction partners, organelle continuity and protein trafficking.

Procedure

1. An initial fluorescence of fluorescent molecules is measured in the region of interest (ROI).
2. The fluorescent molecules are rapidly photobleached by focusing the high-intensity laser beam onto the defined area.
3. The exchange of bleached molecules with unbleached molecules from the surrounding region is followed over time using a low-intensity laser.

Materials

• Optical microscope.
• Light source.
• Fluorescent probe.

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