Fluorescence Decay Statistics

Notice that the Log-Log representation used in the above figure is optional, but is convenient due to the generally large spread of values.

The same histograms as above, built using a linear representation (and using a Decay Statistics Bin of 10) is shown below for comparison:

These histograms can be used to specify another type of pixel rejection criterion than the one defined in the Image Histogram page (Reject Low Count Pixels), as explained next.

The Reject Low Decay Peak Pixels and Reject High Decay Peak Pixels checkboxes, used in combination with the Min Peak Value and Max Peak Value controls, determine which pixel to keep during subsequent analysis, based on the maximum value (or peak value) measured in the fluorescence decay of that pixel.

The Min Values histogram indicates the baseline (minimum of the decay) values within the dataset, and should be considered as a region to avoid for the choice of the Min Peak Value, while the Max Values histogram indicates how many pixels will survive the Reject Low Decay Peak Pixels and Reject High Decay Peak Pixels criteria. For instance, in the example shown above, the minimum decay peak value enforced (500, green cursor) is above the main peak of the "Max Value" histogram, ensuring that only the brightest pixels will be retained and background pixels rejected. The maximum decay peak value (4,000, red cursor) is below the maximum allowed pixel value (4,095) in this dataset, which may help with rejecting saturated pixels. Alternatively, it can be used to reject the brightest pixels in the image, if those are of no use for the analysis.

Finally, the histogram can be used to determine an appropriate peak threshold value to use for the Define ROI(s) above Mean Peak function described in the Using ROIs manual page.

The Reject Low/High Decay Peak Pixels criteria and the Reject Low Count Pixels criterion can be used simultaneously, in which case only pixels meeting all criteria will be retained in subsequent analysis.