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Update Home authored by Vitali Vistunou's avatar Vitali Vistunou
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...@@ -39,7 +39,13 @@ Here we explain how to install and use FCS-calibrated image analysis pipeline. T ...@@ -39,7 +39,13 @@ Here we explain how to install and use FCS-calibrated image analysis pipeline. T
2. Perform three separate sessions of FA analysis: for FP, for POI, and for dye as described [here](https://git.embl.de/grp-ellenberg/FCSAnalyze/-/wikis/Fa_fit_fcs). Do **all** intermediate FA steps (correlations calculation, correction, fitting of correlations with ACF) to obtain mFP.res, POI.res, and dye.res. 2. Perform three separate sessions of FA analysis: for FP, for POI, and for dye as described [here](https://git.embl.de/grp-ellenberg/FCSAnalyze/-/wikis/Fa_fit_fcs). Do **all** intermediate FA steps (correlations calculation, correction, fitting of correlations with ACF) to obtain mFP.res, POI.res, and dye.res.
**Important:** do not forget to save fluctuation traces with ACF fitting curves (Export all traces button) as .cof, .itr files. **Important:** do not forget to save fluctuation traces with ACF fitting curves (Export all traces button) as .cof, .itr files.
3. Prepare the following [structure of files](structure of files). 3. Prepare the following [structure of files](structure of files).
4. Specify parameters in the main user input. You can change any parameters or leave default values. The explanation of every parameter can be founded in a KMIME description menu of the main user input. 4. Specify parameters in the main user input. You can change any parameters or leave default values. Users have several options to calculate an effective confocal volume:
* Using FA session for dye and then specifying the path to dye.res file in the main user input.
* Using [FCSFitM](https://git.embl.de/grp-ellenberg/FCSAnalyze/-/wikis/FCSFitM) provided by Antonio. In this case, users need to specify the volume in the main user input returned by FCSFitM (you can find it in focalVolume.txt). This way **requires the installation of additional software**.
* Other strategies as described in this [article](https://www.picoquant.com/images/uploads/page/files/7351/appnote_quantfcs.pdf). After you received the value of effective confocal volume you can add it into main user input.
The explanation of every parameter can be founded in a KMIME description menu of the main user input.
![input1](uploads/a4c9df72fe2acfc0693cc9b274e0fba6/input1.png) ![input1](uploads/a4c9df72fe2acfc0693cc9b274e0fba6/input1.png)
...@@ -109,4 +115,10 @@ The table with data points in calibration plot (can be used to plot the calibrat ...@@ -109,4 +115,10 @@ The table with data points in calibration plot (can be used to plot the calibrat
6. **map folder** 6. **map folder**
The folder contains processed concentration maps in case the concentration maps metanode was used in analysis. The folder contains processed concentration maps in case the concentration maps metanode was executed.
<br>
In order to add color representation into concentration maps obtained with pipeline user can run 2 commands in Fiji:
* run("16 colors");
* run("Calibration Bar...", "location=[Lower Right] fill=None label=White number=5 decimal=0 font=12 zoom=1 bold overlay");
![f](uploads/cb42b1cfe2a65fed811f9173bec28abe/f.png)
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