Measurement of Diffusion of Mesoscopic Probes in Aqueous Polymer Solutions by Fluorescence recovery after photobleaching (FRAP) (CAT#: STEM-MT-0009-WXH)

Name: Measurement of Diffusion of Mesoscopic Probes in Aqueous Polymer Solutions by Fluorescence recovery after photobleaching (FRAP)
SKU: STEM-MT-0009-WXH
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Introduction

The diffusion of macromolecules through polymer solutions and gels is important in many industrial and biological applications. For example, a better understanding of how macromolecular drugs are transferred in living tissue is essential for the design of macromolecule-based therapies. The transport of macromolecules across membranes, in porous chromatography supports, and in porous media is important to bioseparations. In the food and oil industries, enzymes are often used to degrade polysaccharide chains at very high polymer concentrations. The reactions are diffusioncontrolled, and an understanding of the diffusion limitations is necessary to predict process performance.

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Principle Applications Procedure Materials

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.