Setting Up the PSF Width Tool

To set up the PSF Width Tool:

1. From the softWoRx main menu, select Measure | PSF Width. The PSF Width window is displayed.

2. In the PSF Width window, click the Input button to browse and select the DeltaVision (.dv) file to use for analysis. Alternatively, you can click and drag a .dv file from the Data Folder into this field.

Note: The input to the tool is 10 – 100 frames of spatially well separated beads (smaller than 200nm), or single fluorophores, for the imaging conditions that are used in a localization experiment.

3. To use only a portion of the input image, click the Select Region button and then click and drag the mouse to specify the part of the image you want to analyze. Alternatively, you can use the Details button to define a subset to analyze.

4. Click the Do It button to run the PSF Width tool.

The algorithm looks for well separated events, fits them to a 2D-Gaussian model, and reports the results. Only symmetric, well-spaced beads or single fluorophores are used for measuring the PSF size factor. The results are displayed as a histogram of each fitting event, where the mean of the distribution is the PSF size factor. Additionally, the raw data is displayed with the measured PSF size factor placed next to each bead, or single fluorophore, if it was used to measure the PSF size factor.

Note the PSF size factor which will be used later in the Localization Analysis algorithm.

Note: The PSF size factor is provided in units of pixels, where the size of each pixel is given by the pixel size of the raw data.

Setting Up the PSF Width Tool

1. From the softWoRx main menu, select Measure | PSF Width. The PSF Width window is displayed.

2. In the PSF Width window, click the Input button to browse and select the DeltaVision (*.dv) file to use for analysis. Alternatively, you can click and drag a *.dv file from the Data Folder into this field.

3. To use only a portion of the input image, click the Select Region button and then click and drag the mouse to specify the part of the image you want to analyze. Alternatively, you can use the Details button to define a subset to analyze.

4. Click the Do It button to run the PSF Width tool.

Related Topics

2. In the PSF Width window, click the Input button to browse and select the DeltaVision (.dv) file to use for analysis. Alternatively, you can click and drag a .dv file from the Data Folder into this field.