Frequently Asked Questions

Q: What data format does the program support for image input?
A: MRC files for both 2D electron micrograph and 3D image stack. Signature is able to read TIFF and SPIDER 2D image, which can then be saved to an MRC file for further processing. Notice that the batch processing function supports MRC format only.

Q: What data format does the program support for particle selection output?
A: For 2D coordinates, Signature supports PTK (a proprietary format), PLT (IMAGIC format) and SPD (SPIDER format). As for images, the selected particles can be output to a 3D MRC stack.

Q: How to prepare template images used in the algorithmic particle screening?
A: Template images should be in the form of an MRC stack. They can be created in several ways: 1) 2D projections from a low-resolution density model; 2) class averages from a handful pre-selected particles; or, 3) images cropped from the given micrograph (possibly after low-pass filtering to reduce noise).

Q: How to crop a rectangular section from a 2D image?
A: An image cropping function is provided in 2D-Image OpPanel. Place the mouse cursor at the region of interest, press CTRL, click and drag the mouse left button. Upon mouse button release, the selected section will be displayed in the rectangular monitor in the panel. The cropped image can be saved to an MRC file (the Save button under the monitor). One application of this cropping function is to generate a set of templates directly from a given micrograph and to proceed with automated particle screening.

Q: How to crop a filament segment from a 2D image?
A: When the box by Filament Boxing is checked, image can be cropped along any direction. Place the mouse cursor at the region of interest, press CTRL, click and drag the mouse left button. Upon mouse button release, the selected section will be displayed in the monitor. The image will have been rotated to the horizontal orientation.

Q: A SPIDER particle coordinates file does not match the micrograh display. Why?
A: The problem most likely comes from image conversion: when a micrograph in SPIDER format is converted to the MRC format, the conversion should be processed in the SPIDER program. It is known that, if the IMAGIC program is used, the output image will be flipped. Signature can read the particle selection file originally generated by SPIDER and display it properly on the micrograph, if the image format conversion is correct.