Web : Random Conical Tilt Series Interactive Particle Selection

PURPOSE:

Select marker/image coordinates from a tilted/untilted pair of micrographs (Single exposure random conical tilt series), determine tilt-angle (the sign of the tilt-angle is not determined) and tilt-axis directions in untilted (zero degree) and tilted image.

USAGE:

1. Display untilted (left) image.


2. Display tilted (right) image.


3. Give suffix number for document files that are created / read. If these doc. files don't exist in the current directory they will be created there. If these files already exist they are loaded and particle picking starts with the next particle in the series.


4. If displayed images are same size as the original micrographs which will be used for windowing push: ACCEPT


5. Pick particle location from either untilted (left) or tilted (right) image using left button. Locations are saved in doc files when picked.


6. Pick corresponding particle location from the other image using left button. At least 5 particle pairs have to be selected (seven pairs is preferable). The particles should be distributed throughout the area of the image. A previously selected particle location can be discarded using the right button. The initial particle of a pair can be selected from either image of the pair. When 4-32 particle pairs have been picked the current tilt angle is reported. (This angle is not saved until after running the angle determination on the menu and saving the results.)


7. Use middle button to call menu to:
   
   • Determine tilt and axes rotation angles.

     Tilt angle is found by comparing areas of triangles on both images. The "arealim" parameter from the "Expert menu" sets minimum triangle area used for this comparison.

   • Shift images to view hidden areas.

     Pans images independently.

   • Enhance contrast of images.

     Alters images independently.

   • Pick background window locations from images.

     User sets areas.

   • Erase labels from image.

   Points are not erased from doc files.

   • Draw picked particle locations/numbers on images.
   • Draw fitted locations.
   • Save angles in doc file: dcbxxx

     When satisfied with fitting parameters.

   • Choose expert menu.


8. Once the tilt and axis angles have been calculated you can continue picking particles with predicted locations indicated. After selecting location in either image, the predicted location in other image will be indicated and the user may select the point or adjust its location. At any time a new tilt axis can be determined using all of the selected points.

EXAMPLE OF OPERATION SEQUENCE:

Step.	Notes
Display untilted (left) image
Display tilted image
Pick particle locations	Need at least 5 (better 7) pairs of locations for determining angles and axes.
Determine tilt and axes angles	Can later be run again at any time when more locations are picked.
Repeat	Pick remaining particles
Draw locations.	Can be used any time to display locations.

OUTPUT DOCUMENT FILES:

Name	Contents
dcuxxx	Particle coordinates in untilted image
dctxxx	Particle coordinates in tilted image
dcbxxx	Data common to both images (tilt angle, etc.)

DOCUMENT FILE CONTENTS:

dcuxxx:  (Coordinates in untilted image)

 ; dat/dat  dcu000.dat   Mon Jun 20 09:48:07 2011
 ;             Particle    Reduced-X,Y-location    Original-X,Y-location   
 ;
 0001 6            1          382          214        382          214       1
 0002 6            2           74          339         74          339       1
 0003 6            3          205          207        205          207       1

------------------------------------------------------------------------------

dctxxx:  (Coordinates in tilted image)

 0001 6            1          404          247          404          247     1
 0002 6            2           95          354           95          354     1
 0003 6            3          234          255          234          255     1

------------------------------------------------------------------------------

dcbxxx:  (Data common to the tilt pair)

 ; dat/dat  dcb000.dat   Mon Jun 20 10:30:16 2011

 ;  Key: 121                    #Markers-fitted   #Backgrounds 
 0121 6        0    0    0    0      14             0

 ;  Key: 122   Fitted-flag 
 0122 6          1         

 ;  Key: 123    Untilted-X,Y-Origin      Tilted-X,Y-Origin   Img-Reduction 
 0123 6          382          214      406.683      245.944       1          

 ; Key:  124 Tilt-angle  Untilted-axis-direction-angles (Gamma,Phi) 
 0124 6      50.3053     -78.7234     -77.8619      0    0      0

Subroutines:

fitdoc, pickp, pickmen, fitmen, fitorigin, fitsav, pickdraw, fitdocmen, backmen, spoutfile