AP TOOL - Alignment - Tool for evaluating parameters ||

PURPOSE

Compares a series of experimental images with a series of reference images. For each experimental image, it finds the in-plane Euler rotation angle, and X, Y translational shifts which align the image with the most-similar reference image.

SEE ALSO

AP SH	[Alignment - multi-reference, exhaustive rotation & shift ||*]
AP SHC	[Alignment - multi-reference, fast exhaustive rotation & shift ||]

USAGE

.OPERATION: AP TOOL

.EVALUATE SHIFT OR RINGS (SH/RI): SHIFTS

.TEMPLATE FOR REFERENCE IMAGES: REF***
[Give the template for the name of the existing file series of 2D reference images (typically projections).]

.FILE NUMBERS OR SELECTION DOC. FILE NAME: SELECTREF
[Enter numbers of reference files. The file numbers can also be read from a selection document file where file numbers are contained in the first register (not the keys).]

If evaluating shifts the following prompt occurs:

.MAX TRANSLATION SEARCH RANGE: 14
[The search for translation parameters will be restricted to +/- search range, performed every "step size" pixel.

Otherwise if evaluating Rings the following prompt occurs:

.TRANSLATION SEARCH RANGE: 14
[The search for translation parameters will be restricted to +/- search range, performed every "step size" pixel.

If evaluating shifts the following prompt occurs:

.FIRST, LAST RING, RING STEP & RAY STEP: 5,176,1,1
[The search for rotational alignment will be restricted to pixels with radii in the specified range (Here: 5-176), performed every 'ring step' radius and on every 'ray step' radial ray.

Otherwise if evaluating rings the following prompt occurs:

.FIRST, LAST RING: 5,176
[The search for rotational alignment will be restricted to pixels with radii in the specified range (Here: 5-176).

.REFERENCE IMAGES ANGLES DOCUMENT FILE: Refangles
[Enter the name of the document file containing Eulerian angles for the reference images: Psi, Theta, Phi.]

.TEMPLATE FOR IMAGE SERIES TO BE ALIGNED: DAT***
[Give the template name of the existing file series of experimental images. These images will be checked for alignment versus the reference images.]

.FILE NUMBERS OR SELECTION DOC. FILE NAME: 1-2100
[Enter numbers of experimental files. The file numbers can also be read from a selection document file where file numbers are contained in the first register (not the keys).]

.CHECK MIRRORED POSITIONS (0=NOCHECK / 1=CHECK)?: 1
[Optional check of the mirrored reference image. By using this check and only providing reference images from one hemisphere of projection directions, speed can be doubled. See note: 1 below.

.OUTPUT ALIGNMENT DOCUMENT FILE: PARM101
[This is the only output produced by this operation. (Will append to existing output file of same name). It contains 15 register columns. See 'AP SH' for details.

NOTES

1. The main take-away-message from experimentation with use of this operation is that the alignment operation does not need the 'best' match to give decent reconstructions. Due to the high levels of noise common in cryo-em images, reconstruction is a 'statistical' summation and sub-maximal alignments combine to still give decent reconstructions.

2. Explanation of output for shift analysis.

   SHIFT RANGE	STEP VALUE	#BEST FOUND	%BEST FOUND	%BEST CC	%ON BORDER	TIME
   Shift range	Step size	Number of best CC matches found	% of best CC matches found	% of best CC	% of best CC matches occurring on border of shift range	Relative run time

3. Explanation of location of maximum matches
   Number of times best match for each expermental images occurs at each shift location.

4. Explanation of output for ring skip analysis.

   SKIP AMOUNT	#BEST FOUND	%BEST FOUND	%BEST CC	RELATIVE TIME
   Skip value	Number of best CC matches found	% of best CC matches found	% of best CC	Relative run time

5. Output document file produced by this operation contains 15 register columns. The contents are the same as operation: 'AP SH' except for the following:
   • Cumulative X shift.
     This is the sum of any shift from the 'experimental images align. document file' and any current shift.
     The shifts reported here have NOT been adjusted to compensate for any rotation, so that they are not appropriate for use with SPIDER operations such as 'RT SQ'.
   • Cumulative Y shift.
     This is the sum of any shift from the 'experimental images align. document file' and any current shift.
     The shifts reported here have NOT been adjusted to compensate for any rotation, so that they are not appropriate for use with SPIDER operations such as 'RT SQ'.
   • Current X shift.
     X shift necessary to align the experimental image with the current reference projection. The shifts reported here have NOT been adjusted to compensate for any rotation, so that they are not appropriate for use with SPIDER operations such as 'RT SQ'.
   • Current Y shift.
     Y shift necessary to align the experimental image with the current reference projection.
   The shifts reported here have NOT been adjusted to compensate for any rotation, so that they are not appropriate for use with SPIDER operations such as 'RT SQ'.

SUBROUTINES

APSH_TOOL, APRINGS, NORMASC, CROSRNG_2, FRNGS, AP_STAT, RINGWE, APPLYWS

CALLER

UTIL4