Parameter importance in FRAP acquisition and analysis: a simulation approach.

Fluorescence recovery after photobleaching (FRAP) is a widespread technique used to determine intracellular reaction and diffusion parameters. In recent years, due to technical advances and an increasing number of mathematical models for analysis, there was a resurging interest in FRAP applications. However, care has to be taken when inverting parameters from such data. We study potential influences on FRAP acquisition and analysis like initial fluorescence distribution, membrane passage, and geometrical aspects. Monte Carlo simulations are employed for the investigation of reaction-diffusion processes to additionally include cases in which no analytical description is available. To assess the importance of influencing factors we apply a sensitivity method based on elementary effects providing an estimate for the global parameter space. The combination of simulations and sensitivity measure helps us to predict ranges of parameters used in acquisition and analysis for which a reliably inversion of reaction-diffusion parameters is possible. Using this approach, we show that FRAP data are highly susceptible to misinterpretation. However, by identifying the parameters of susceptibility, our analysis provides the means for taking measures to significantly improve FRAP data interpretation and analysis.