Daybook Analysis – Version 2.0.0 – October 2024
The intensity response of a fluorescence microscope expresses the output digital signal to an input photon flux. It depends on many aspects of the microscope, both in the illumination path and the detection path.
For any fluorescence microscope, the overall intensity response may evolve over time. For example, variations in the illumination power, detector sensitivity, optical alignment, etc. can lead to intensity response fluctuations. It is therefore important to know the intensity response when intensity quantification in images of a biological sample, acquired at different times, is being performed.
The “intensity response” analysis provides the intensity response of the imaging system to 16 intensity levels following a linear evolution, as well as quantitative metrics such as intensity values and pattern dynamic range. Monitoring these metrics, reveals the evolution of the intensity response over time, with respect to reference values.
The “intensity response” analysis is associated with the “4×4 intensity gradation” (Pattern family C) and “2×16 intensity gradation” (Pattern family D) patterns (see Figure 1).
Figure 1: Example images of the “intensity gradation” patterns, fulfilling the acquisition recommendations. Left: “4×4 intensity gradation” pattern. Right: “2×16 intensity gradation” pattern.