title: Fluorescence recovery after photobleaching in material and life sciences: putting theory into practice.
abstract: Fluorescence recovery after photobleaching (FRAP) is a versatile tool for determining diffusion and interaction/binding properties in biological and material sciences. An understanding of the mechanisms controlling the diffusion requires a deep understanding of structure-interaction-diffusion relationships. In cell biology, for instance, this applies to the movement of proteins and lipids in the plasma membrane, cytoplasm and nucleus. In industrial applications related to pharmaceutics, foods, textiles, hygiene products and cosmetics, the diffusion of solutes and solvent molecules contributes strongly to the properties and functionality of the final product. All these systems are heterogeneous, and accurate quantification of the mass transport processes at the local level is therefore essential to the understanding of the properties of soft (bio)materials. FRAP is a commonly used fluorescence microscopy-based technique to determine local molecular transport at the micrometer scale. A brief high-intensity laser pulse is locally applied to the sample, causing substantial photobleaching of the fluorescent molecules within the illuminated area. This causes a local concentration gradient of fluorescent molecules, leading to diffusional influx of intact fluorophores from the local surroundings into the bleached area. Quantitative information on the molecular transport can be extracted from the time evolution of the fluorescence recovery in the bleached area using a suitable model. A multitude of FRAP models has been developed over the years, each based on specific assumptions. This makes it challenging for the non-specialist to decide which model is best suited for a particular application. Furthermore, there are many subtleties in performing accurate FRAP experiments. For these reasons, this review aims to provide an extensive tutorial covering the essential theoretical and practical aspects so as to enable accurate quantitative FRAP experiments for molecular transport measurements in soft (bio)materials.
authors: SP Food and Bioscience,PO 5401, SE-402 29, Göteborg,Sweden.
authors: Niklas
authors: N
authors: Lorén
authors: SP Food and Bioscience,PO 5401, SE-402 29, Göteborg,Sweden.
authors: Joel
authors: J
authors: Hagman
authors: Department of Mathematical Sciences,Chalmers University of Technology,SE-412 96 Göteborg,Sweden.
authors: Jenny K
authors: JK
authors: Jonasson
authors: Biophotonic Imaging Group,Laboratory of General Biochemistry and Physical Pharmacy,Ghent University,9000 Ghent,Belgium.
authors: Hendrik
authors: H
authors: Deschout
authors: Department of Chemical and Biological Engineering,Chalmers University of Technology,SE-412 96 Göteborg,Sweden.
authors: Diana
authors: D
authors: Bernin
authors: Nanophysics Department,Istituto Italiano di Tecnologia,Via Morego 30, 16163 Genova,Italy.
authors: Francesca
authors: F
authors: Cella-Zanacchi
authors: Nanophysics Department,Istituto Italiano di Tecnologia,Via Morego 30, 16163 Genova,Italy.
authors: Alberto
authors: A
authors: Diaspro
authors: Institute for Soft Matter and Functional Materials, Helmholtz Center Berlin,12489 Berlin,Germany.
authors: James G
authors: JG
authors: McNally
authors: Hasselt University,Campus Diepenbeek,Martelarenlaan 42,3500 Hasselt,Belgium.
authors: Marcel
authors: M
authors: Ameloot
authors: Hasselt University,Campus Diepenbeek,Martelarenlaan 42,3500 Hasselt,Belgium.
authors: Nick
authors: N
authors: Smisdom
authors: Ian Wark Research Institute,University of South Australia,Adelaide,Australia.
authors: Magnus
authors: M
authors: Nydén
authors: SP Food and Bioscience,PO 5401, SE-402 29, Göteborg,Sweden.
authors: Anne-Marie
authors: AM
authors: Hermansson
authors: Department of Mathematical Sciences,Chalmers University of Technology,SE-412 96 Göteborg,Sweden.
authors: Mats
authors: M
authors: Rudemo
authors: Biophotonic Imaging Group,Laboratory of General Biochemistry and Physical Pharmacy,Ghent University,9000 Ghent,Belgium.
authors: Kevin
authors: K
authors: Braeckmans
mesh_terms: D005453:Fluorescence; D038761:Photobleaching