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- Go to the Launch Pad and click the N-STORM Analysis button to open an empty N-STROM Analysis window. In the new window click on the folder icon, locate the file you want to process and click Open to display the STORM dataset.
- The widefield images of all frames you have acquired will be displayed. Use the slider at the bottom of the image window to navigate through the different time points. You can right click on the slider at any point to open a menu that allows you to set a start and/or an end point for the image processing, e.g. this can be useful if the first few frames are too bright to localize any molecules and you therefore do not want to include them in the analysis.
- Press the Peak Statistics button on the right and hover over the background to get an idea of the background intensity in your sample. Do this for a few frames and note down the average value. Note that the peak statistics option is not available when Auto Minimum Height is selected in the Identification Settings.
- Click on the Identification Settings icon (or the STORM Analysis icon and then Identification settings) to go to the settings menu and configure the Minimum Height with the value that you measured in the previous step. Alternatively tick the Auto Minimum Height checkbox to allow the software to determine the minimum intensity value for the image processing. The Maximum Height is automatically set to 65,536, which corresponds to the number of grey levels of the 16-bit EMCCD camera is used for the imaging. The CCD Baseline value is 100. Select 3D if it is a 3D dataset otherwise leave the checkbox unticked. By clicking the double arrow button you get to an advanced menu where you can adjust further Identification settings like the Minimum Width (i.e. 200) and the 2D or 3D Maximum Width (i.e. 400 or 700 respectively). Set the Fit Width (nm) to 300, the Max. Axial Ratio to 1.3 for 2D data and 2.5 for 3D data and the Max. Displacement (pix) to 1. Click OK after you have set up all Identification Settings.