Difference: Jeol1230Instructions (27 vs. 28)

Revision 2820 Feb 2008 - Main.BillRice

 
META TOPICPARENT name="CryoEM"
Contents

JEOL 1230 Instructions

Specimen Insertion

  • Flip toggle switch to "pump" (G)
  • Line up pin on holder with goniometer(G)
  • Insert specimen holder to first sample stop (marked line indicates correct depth)
    • You will hear a click
    • Orange light will go on
  • Monitor Vacuum condition:
    • Go to "FR Panel" "monitor" "vacuum conditions"
    • When SPC value is <50 insert holder by turning the holder slowly clockwise 1/8 turn to the 1st stop
  • Slowly allow the vacuum to pull the holder in
  • Grasp holder firmly then turn the holder slowly clockwise 1/4 turn to 2nd stop
  • Slowly allow the vacuum to pull the holder in
  • Wait for green light to come on before turning on beam
    sampleinsertion.jpg

Aquiring a Beam

  • Open "HT" Menu (top left screen)
  • Select beam "on" *Filament current should be around 70%
  • Be patient, it takes about one minute for the beam to appear

Condenser Aperture Alignment

  • At 5000x
  • Use "Brightness" knob to condense beam on function box(F)
  • press "Beam Shift" (F)
  • Right click mouse button and drag the beam to central position
  • Spread Beam (over clockwise) with "Brightness" knob (F)
  • If it does not spread evenly, center with the condenser aperture x-y controls
  • condense beam with "Brightness" (F)
  • If beam is not centered, repeat
  • The beam should expand/ contract concentrically around the beam centre as the brightness is adjusted

Objective Aperture Alignment

  • Select Diffraction mode by pressing "Diffraction" (F)
  • Set the camera length to 150cm
  • Condense beam with "Brightness" (F)
  • Focus beam with "Diff focus" (F)
  • Insert objective aperture
  • Center aperture with x-y controls

Z Height Adjustment

  • On the menu, go to Monitor, then enter Lens condition sub-menu.
  • By adjusting the focus, set the objective lens current to 4.7.
  • Enter Z cont mode, then click AUTO to set specimen eucentric height.

Diffraction

  • Select Diffraction button (F)
  • Use Align Adjust for Diffraction shift
  • PL Align

Low Dose

  • "Photo" menu
  • "MDS"

Specimen Removal

  • Switch the beam is off
  • Switch the toggle to "Air" (G)
  • Monitor Vacuum condition:
    • Go to "FR Panel" "monitor" "vacuum conditions"
  • Pull out specimen holder to 1st stop
  • Rotate anti-clockwise 1/4 turn
  • Pull out the specimen holder to the 2nd stop
  • Rotate anti-clockwise 1/8 turn
  • Wait for the airlock "SPC" to vent
  • Gently remove the specimen rod (G)
    • If the use of force is necessary the SPC is not vented

1230 ShutDown Checklist

FilamentExchange

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SerialEM Operation

Startup

  1. Launch Digital Micrograph
  2. Launch Temserver20xx.exe
  3. Launch your copy of SerialEM

Montaging

  • To do a low-mag montage, go to 100X or 80X, fully expand beam, take a test Record image to be sure images are OK
  • Go to Navigator menu, choose to Open
  • Go to Navigator menu, choose Setup Full Montage
  • Choose to save all header info, save file as FILENAME.st
  • Increase overlap percent to 20 (3X3 images for full montage at 100X)
  • Click Start on montage menu
  • Montage will begin. If it appears to get stuck, quickly toggle mags
  • When done, click YES to make a new map
  • Try checking and unchecking Treat as very sloppy montage -- sometimes it is better, sometimes worse
  • Go to File menu, click close to close the file for writing (do not want to add to this file)

Alignment of Montages

  • First map is registration 1
  • Make a new montage, either at a different mag or at a different grid orientation
  • Follow instructions above, but before you click start, CHANGE CURRENT REGISTRATION TO 2
  • Make the new map -- you now have old map in registration 1 and new in registration 2
  • Need at least 3 points to align the maps (determine a shift/rotation matrix)
  • Choose three obvious points, click Add Points, and add points. When done, click stop
  • Mark each point as a Registration point, numbered 1, 2 and 3
  • Re-open first map by double clicking
  • CHANGE CURRENT REGISTRATION BACK TO 1
  • Click Add Points, and add the three corresponding points on the old map
  • Mark these as Registration Points , again 1,2,3, IN THE SAME ORDER AS ON MAP 2
  • Change current registration back to 2, and re-load new map
  • Under Navigator menu, choose Transform items
  • Map 1 is now transformed onto map 2
 * Set ALLOWTOPICVIEW =

-- KdDerr - 13 Jun 2006

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