IRF stands for Instrument Response Function, and it reflects how a component with an infinitive short lifetime is seen by the instrument. Of course, all components of the instrument contribute to both shape and width of the IRF. This width is an indicator the timing resolution of the instrument.
The IRF width is approx. a quadratic sum of the widths of all contributions. The observed decay in a time domain measurement is a convolution of the 'real' decay with the IRF. Therefore for accurate measurements of lifetimes, especially near to the timing resolution, the instrument response function (IRF) should be taken into account. For IRF Measurements, several strategies are possible which are described here: https://www.tcspc.com/doku.php/howto:how_to_measure_the_instrument_response_function_irf
For the further analysis it is assumed that:
For IRF Measurements, several strategies are possible which are described here.
1. First the measured IRF needs to be made accessible for being used in the analysis. Therefore, highlight the file with the measured IRF (single mouse click on the .ptu-file)
2. In Analysis tab, under TCSPC, find `TCSPC Decay`. Click Start;
Response: you see this kind of curve:
(Before selecting a detection channel the peak is above 100000, so fine for this step)
3. Select the proper detection channel (1 or 2) and Click”Calculate Decay”. The decay is then updated.
4. Select the decay range by Setting a `Time Gate` (move the red bars) to limit the decay the area where this has an impact.
Setting a time gate removes the noise, and the fit curves usually become smoother.
5. Then click again on `Calculate Decay`. As a result you will see:
Select the linear range for measured curve (on top right), right click, activate `Data Reader`, and try to find FWHM and see how broad it is. 2)
In this example channel 2 was selected, now the data can be “saved as IRF”, and be imported for future analysis accordingly. Please check the different FLIM-related tutorials for more information about the fitting.