Scientific Volume Imaging

The Huygens Bleaching Corrector is a tool that allows you to track and adjust for fading of the fluorescent signal over time or in the z direction. By correcting the images you can perform more reliable image analysis on slices or frames of the image or time-series.

Any fluorescence microscopy technique in which the sample is imaged over time (Widefield, Spinning Disc, confocal or STED time series), can be affected by sample bleaching. Also imaging over multiple planes could result in bleaching of deeper planes. Bleaching is caused by the continued or repeated excitation of the fluorescent molecules causing them to emit less light leading to fading of the image. Bleaching can be caused by photo-oxidation, organic reactions and multi-photon events, also see Bleaching-Effects

Bleaching depends on the fluorescent molecule used and the environment of the sample. Bleaching can be reduced by imaging over a shorter time, using less intervals or reducing the excitation laser power. If these settings cannot be altered, bleaching can be reduced by using the more photostable fluorescent molecules or by deoxygenating the sample. With Huygens deconvolution you can resolve high resolution images even with low signal. Therefore, you can design your experiment with lower acquisition times or laser power, which helps to prevent bleaching. If bleaching has already occured in your image, you can use the Huygens Bleaching Corrector to detect and correct for by the Huygens Bleaching Corrector.

When performing time-series imaging there is always a risk of bleaching. Bleaching can complicate any data analysis. If you wish to track objects or determine colocalization bleaching should be corrected for, not doing so might result in unreliable data.