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writingroom:lifetime-fitting_using_the_flim_script [2020/05/12 14:18] – ruckelshausenhowto:lifetime-fitting_using_the_flim_script [2020/05/12 14:35] – ↷ Links adapted because of a move operation ruckelshausen
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 **Response:​**\\ ​ **Response:​**\\ ​
 The file ''Kidney_Cells_FLIM.ptu'' is opened with the FLIM analysis. Thereby, a new Window opens:  The file ''Kidney_Cells_FLIM.ptu'' is opened with the FLIM analysis. Thereby, a new Window opens: 
-{{ :writingroom:kidneyfullscreen3.png |}}+{{ howto:kidneyfullscreen3.png |}}
  
 **Note:**\\ **Note:**\\
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   * Open the "Region of Interest" drop-down menu, tick "Use ROI" checkbox and click "ROI from threshold".\\   * Open the "Region of Interest" drop-down menu, tick "Use ROI" checkbox and click "ROI from threshold".\\
  
-{{ :writingroom:activateroi.png |}}+{{ howto:activateroi.png |}}
  
 **Response:​**\\ **Response:​**\\
 A large window with three sections will appear, where you can set a threshold for an image analysis visually.   A large window with three sections will appear, where you can set a threshold for an image analysis visually.  
  
-{{ :writingroom:selectthreshold_kidney3.png |}} +{{ howto:selectthreshold_kidney3.png |}} 
  
 **Note:**\\ **Note:**\\
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-{{ :writingroom:roi0_kidney3.png |}}+{{ howto:roi0_kidney3.png |}}
  
    
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   * In the fourth box, "Decay Fitting", below the image, select the Fitting Model drop-down list select "n-Exponential Reconvolution Fit".\\   * In the fourth box, "Decay Fitting", below the image, select the Fitting Model drop-down list select "n-Exponential Reconvolution Fit".\\
  
-{{ :writingroom:nexpon_fitmodel.png?400 |}}+{{ howto:nexpon_fitmodel.png?400 |}}
  
 ** Response:**\\ ** Response:**\\
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   * From "Decay" drop-down list select ''ROI 0'' and click "Initial Fit" (marked in orange).\\   * From "Decay" drop-down list select ''ROI 0'' and click "Initial Fit" (marked in orange).\\
-{{ :writingroom:monoexpo_roi0_kidney_3.png |}}+{{ howto:monoexpo_roi0_kidney_3.png |}}
    
 ** Response:**\\  ** Response:**\\ 
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   * In this example it is obvious that the applied single exponential reconvolution fit is not suited to fit the decay curve. Increase the "Model Parameters" to n = 2. Again, click on "Initial fit".\\   * In this example it is obvious that the applied single exponential reconvolution fit is not suited to fit the decay curve. Increase the "Model Parameters" to n = 2. Again, click on "Initial fit".\\
  
-{{ :writingroom:biexpo_roi0_kidney_3.png |}}+{{ howto:biexpo_roi0_kidney_3.png |}}
  
 ** Response:**\\ ** Response:**\\
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-{{ :writingroom:3expo_roi0_kidney_3.png |}}+{{ howto:3expo_roi0_kidney_3.png |}}
  
  
   * Three components exponential decay seems to be suitable for this ROI. However we can try four exponential decay and see the result:\\   * Three components exponential decay seems to be suitable for this ROI. However we can try four exponential decay and see the result:\\
  
-{{ :writingroom:4expo_roi0_kidney_3.png |}}+{{ howto:4expo_roi0_kidney_3.png |}}
  
 **Response:**\\ **Response:**\\
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-{{ :writingroom:limitamplitude_2.png?600 |}}+{{ howto:limitamplitude_2.png?600 |}}
    
  
   * Set the same limit for A2 and A3.\\   * Set the same limit for A2 and A3.\\
   * Uncheck the boxes for the individual lifetime components, Bkgr(Decay), Bkgr(IRF) and Shift(IRF) in order to fix these values, IRF and Background. Then in the pixel-by-pixel fit, these values are kept constant. Do NOT click "Initial Fit" or "Fit" at this stage!\\   * Uncheck the boxes for the individual lifetime components, Bkgr(Decay), Bkgr(IRF) and Shift(IRF) in order to fix these values, IRF and Background. Then in the pixel-by-pixel fit, these values are kept constant. Do NOT click "Initial Fit" or "Fit" at this stage!\\
-{{ :writingroom:parameterfix_roi0_kidney_2.png?400 |}}+{{ howto:parameterfix_roi0_kidney_2.png?400 |}}
  
   * Click "FLIM Fit" in the upper panel.\\   * Click "FLIM Fit" in the upper panel.\\
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   * An updated lifetime frequency histogram on the upper right. The individual lifetime components are displayed as vertical bars, as they are not varied in the pixel-by-pixel fit.\\   * An updated lifetime frequency histogram on the upper right. The individual lifetime components are displayed as vertical bars, as they are not varied in the pixel-by-pixel fit.\\
  
-{{ :writingroom:flimfit_kidney_2.png |}}+{{ howto:flimfit_kidney_2.png |}}
    
   * The three lifetime components can be displayed in a false color scale. Therefore switch the display to "RGB" (above the intensity and color scale) and adapt the intensities of each color as preferred.\\   * The three lifetime components can be displayed in a false color scale. Therefore switch the display to "RGB" (above the intensity and color scale) and adapt the intensities of each color as preferred.\\
-{{ :writingroom:rgb_flimfit_kidney_2.png |}}+{{ howto:rgb_flimfit_kidney_2.png |}}
  
 **Note:**\\ **Note:**\\
howto/lifetime-fitting_using_the_flim_script.txt · Last modified: 2023/08/03 14:16 by lan