Förster Resonance Energy Transfer Measurements in Living Plant Cells

Schmidtpott S, Seidel T (2021)
Journal of visualized experiments : JoVE (172).

Zeitschriftenaufsatz | Veröffentlicht | Englisch
 
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Abstract / Bemerkung
Sensitized emission-based Forster resonance energy transfer (FRET) experiments are easily done but depend on the microscopic setup. Confocal laser scanning microscopes have become a workhorse for biologists. Commercial systems offer high flexibility in laser power adjustment and detector sensitivity and often combine different detectors to obtain the perfect image. However, the comparison of intensity-based data from different experiments and setups is often impossible due to this flexibility. Biologist-friendly procedures are of advantage and allow for simple and reliable adjustment of laser and detector settings. Furthermore, as FRET experiments in living cells are affected by the variability in protein expression and donor-acceptor ratios, protein expression levels must be considered for data evaluation. Described here is a simple protocol for reliable and reproducible FRET measurements, including routines for the estimation of protein expression and adjustment of laser intensity and detector settings. Data evaluation will be performed by calibration with a fluorophore fusion of known FRET efficiency. To improve simplicity, correction factors have been compared that have been obtained in cells and by measuring recombinant fluorescent proteins.
Erscheinungsjahr
2021
Zeitschriftentitel
Journal of visualized experiments : JoVE
Ausgabe
172
eISSN
1940-087X
Page URI
https://pub.uni-bielefeld.de/record/2956415

Zitieren

Schmidtpott S, Seidel T. Förster Resonance Energy Transfer Measurements in Living Plant Cells. Journal of visualized experiments : JoVE. 2021;(172).
Schmidtpott, S., & Seidel, T. (2021). Förster Resonance Energy Transfer Measurements in Living Plant Cells. Journal of visualized experiments : JoVE(172). https://doi.org/10.3791/62758
Schmidtpott, S., and Seidel, T. (2021). Förster Resonance Energy Transfer Measurements in Living Plant Cells. Journal of visualized experiments : JoVE.
Schmidtpott, S., & Seidel, T., 2021. Förster Resonance Energy Transfer Measurements in Living Plant Cells. Journal of visualized experiments : JoVE, (172).
S. Schmidtpott and T. Seidel, “Förster Resonance Energy Transfer Measurements in Living Plant Cells”, Journal of visualized experiments : JoVE, 2021.
Schmidtpott, S., Seidel, T.: Förster Resonance Energy Transfer Measurements in Living Plant Cells. Journal of visualized experiments : JoVE. (2021).
Schmidtpott, Sonja, and Seidel, Thorsten. “Förster Resonance Energy Transfer Measurements in Living Plant Cells”. Journal of visualized experiments : JoVE 172 (2021).

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PMID: 34251368
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