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--- howto:calculate_ratiometric_fret-images [2014/05/23 07:41] – admin
+++ howto:calculate_ratiometric_fret-images [2014/06/18 09:20] – admin
@@ Line 11: / Line 11: @@
 This tutorial shows step-by-step, how the "FRET Image" script of [[products:SymPhoTime64|SymPhoTime 64]] can be used to calculate pixel-by-pixel the average [[glossary:fret|FRET]] efficiency in an image containing several cells transfected with Cerulean/YFP constructs. To calculate the average FRET efficiency, a ratiometric approach based on the intensities in the donor and the acceptor channel is used.
-**Note:** [[glossary:fret|FRET]] is a method to determine molecular interactions or conformational changes based on a suited donor/acceptor pair.\\
+===== Background Information =====
+[[glossary:fret|FRET]] is a method to determine molecular interactions or conformational changes based on a suited donor/acceptor pair.\\
 Intensity FRET as used here can only be used to calculate average FRET efficiencies. In case of a mixed population, these populations can only be separated using a donor dye with a single exponential decay kinetic and analyzed with lifetime data. This is not the scope of this tutorial.\\
 The script usage is demonstrated on an image of several cells which have been transfected with CENP-B-Cerulean and YFP-CENP-A. Cerulean is a CFP variant an frequently used as a FRET donor. The CENP-complex consists on several proteins and is involved in mitosis. For further details on the sample, see S. Orthaus et al., Chembiochem. 2008, 9, 77-92, and the PicoQuant application note [[http://www.picoquant.com/images/uploads/page/