Fluorescence microscopty - Fluorescence contribution factors

4 important questions on Fluorescence microscopty - Fluorescence contribution factors

For what is photobleaching the biggest problem?

Live cell imaging, because of the lifetime of fluorescent proteins like GFP.

How is bleaching minimized?

- Choosing low excitation intensities and low exposure times (trade-off with signal level)

In contrast to bleaching, quenching...

Does not damage the fluorophore but it does lower the signal.
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5 properties that affect light intensity of detected fluorophore light:

1. Light source.
2. Filters
3. Lenses
4. Specimen
5. Detector

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