Update Home authored by Vitali Vistunou's avatar Vitali Vistunou
### Welcome to a Wiki page of FCS pipeline! ### Welcome to a Wiki page of FCS pipeline!
Here we explain how to install and use Fluorescence Correlation Spectroscopy (FCS) pipeline. The FCS pipeline is an FCS-based calibrated image analysis software for the interactive and automated processing of FCS data. Here we explain how to install and use Fluorescence Correlation Spectroscopy (FCS) pipeline. The FCSpipeline is an FCS-based calibrated image analysis software for the interactive and automated processing of FCS data.
#### 1. [Installation](installation) #### 1. [Installation](installation)
#### 2. [Structure of workflow](#structure-of-workflow) #### 2. [Structure of workflow](#structure-of-workflow)
...@@ -10,7 +10,7 @@ Here we explain how to install and use Fluorescence Correlation Spectroscopy (FC ...@@ -10,7 +10,7 @@ Here we explain how to install and use Fluorescence Correlation Spectroscopy (FC
- three user inputs: - three user inputs:
- main user input (general parameters) - main user input (general parameters)
- plot parameters input (customizing your plot and setting the QC parameters) - plot parameters input (customizing your plot and setting the QC parameters)
- WT user input (calculating the background offset for correction step in FA) - WT user input (calculating the background offset for correction step in Fluctuation Analyzer (FA))
- loading metanodes: - loading metanodes:
- collecting fluctuation data (FP&POI, dye metanode) - collecting fluctuation data (FP&POI, dye metanode)
- loading images and coordinates (FP&POI&WT images metanode) - loading images and coordinates (FP&POI&WT images metanode)
...@@ -19,11 +19,11 @@ Here we explain how to install and use Fluorescence Correlation Spectroscopy (FC ...@@ -19,11 +19,11 @@ Here we explain how to install and use Fluorescence Correlation Spectroscopy (FC
- visualization nodes - visualization nodes
#### Procedure #### Procedure
1. This step can be omitted. Process a WT data with Fluctuation Analyzer (FA) to obtain WT.res (only loading data and saving data steps are needed). In FCSPipeline KNIME workflow, fill a WT user input and execute Python Source node. Use returned offset value for correction steps in further FA sessions for FP and POI. 1. This step can be omitted. Process a WT data with FA to obtain WT.res (only loading data and saving data steps are needed). In FCSPipeline KNIME workflow, fill a WT user input and execute Python Source node. Use returned offset value for correction steps in further FA sessions for FP and POI.
2. Perform two separate sessions of FA analysis: for FP and for POI. Do all intermediate FA steps (correlations calculation, correction, fitting of correlations with ACF) to obtain mFP.res and POI.res. **Important:** do not forget to save fluctuation traces with ACF fitting curves (Export all traces button) as .cof, .itr files. 2. Perform two separate sessions of FA analysis: for FP and for POI. Do all intermediate FA steps (correlations calculation, correction, fitting of correlations with ACF) to obtain mFP.res and POI.res. **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 a main user input 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 description menu of KNIME node. > You can change any parameters or leave default values. The explanation of every parameter can be founded in a description menu of the KNIME node.
![input1](uploads/a4c9df72fe2acfc0693cc9b274e0fba6/input1.png) ![input1](uploads/a4c9df72fe2acfc0693cc9b274e0fba6/input1.png)
...@@ -58,14 +58,18 @@ info.csv is generated inside the main user input. This is the main output with a ...@@ -58,14 +58,18 @@ info.csv is generated inside the main user input. This is the main output with a
info.csv includes: info.csv includes:
* diffusion coefficient of dye and confocal volume * diffusion coefficient of dye and confocal volume
* the names of directories and files with FCS data * the names of directories and files that are used in FCSpipeline
* path to the main directory * path to the main directory
* calibration plot parameters and its errors * calibration plot parameters and its errors
2. **calibration.csv** 2. **calibration.csv**
Here you can find the concentrations of POI calculated using calibration plot. Here you can find the concentrations of POI calculated using calibration parameters.
3. **calibration_plot.png** 3. **calibration_plot.png**
The image of the final calibration plot. The image of the final calibration plot.
4. **calibration_plot.csv**
The table with data points in calibration plot (can be used to plot the calibration plot with other software)