I'm consistently having problems with this and haven't figured out how to solve it.

Are there good approaches to avoiding bleaching in the first instance?

Is creation of a photobleach curve appropriate? And if so how do I go about doing this? Is a photobleach curve created from a set of calibration experiments representative enough to be used to correct for all subsequent experiments (assuming the microscope settings are kept consistent)? Or do I need to create a separate photobleach curve for each time-lapse in a different ROI to that which I will be quantifying my experimental data from?

Is there a post-capture method of correcting for bleaching? I have heard about the photobleach correction in imageJ (Fiji) however I'm not sure if this counts as manipulation of data and not good practice for subsequent quantification.

Many thanks!