Jose Miguel de la Rosa Trevin edited this page May 21, 2016 · 2 revisions

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spider - align ap sr

This protocol provides the wrapper for the reference-free alignment algorithm in SPIDER. It is based on AP SR program and performs both translational and rotational alignment.

The main part of the procedure brings two objects into register. This is achieved by alternating shift and rotational alignment until the relative change of position is smaller than 0.5 pixels. After alignment of all the objects (which constitutes one iteration step of the procedure) the global average is centered using the low-pass filtered disc of the same size as the object and alignment parameters for all the images are modified accordingly. Iterations are repeated until stable alignment is reached (see above).

Input parameters

There are only few parameters to choose (fig. 1). Input particles should have positive contrast, i.e. objects should be bright on the dark background. You have to decide the limits of integration for the rotational search (in terms of inside and outside rings). The outer radius has to correspond to the particle size. To avoid oversampling (and thus the relatively large error of interpolation) an inner radius of 5 px was found to be sufficient for all practical purposes.

Figure 1. GUI input form of the spider - align ap sr protocol

Analyzing results

When the protocol is finished you may click on the Analyze Results button. The produced output are the final total average and a stack of aligned particles (Fig. 2).

02.SpiderProtAlignAPSR Results
Figure 2. Displaying alignment results


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