MSI set up in more details » History » Version 26
Anchi Cheng, 08/16/2010 07:32 PM
1 | 1 | Amber Herold | h1. MSI set-up in more details |
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4 | 10 | Amber Herold | This section shows some of the more important and complicated steps in the set-up checklist. It assumes that the user knows the nomenclatures. See the full documentation if you want the detailed explanation. It also assumes that general set-up and calibrations exist from previous sessions so that most calibrations can remain unchanged. The steps that are NRAMM specific are marked. MSI now features stage Eucentric height correction at each square. |
5 | 1 | Amber Herold | |
6 | 10 | Amber Herold | NOTE: The Tietz imaging software (EMMenu) must be turned OFF before Leginon is started. In contrast to this, the Gatan imaging software (Digital Micrograph) must be ON before Leginon is started. |
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10 | 10 | Amber Herold | h2. Starting Up |
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14 | 10 | Amber Herold | Set up is most stable with a good microscope alignment. |
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16 | 11 | Amber Herold | 1. Initial alignment at the microscope See Chapter on [[Microscope Set-up|Microscope setup]] for details. |
17 | 1 | Amber Herold | |
18 | 10 | Amber Herold | * scope> Insert specimen holder |
19 | * scope> Put stage to eucenter and reset defocus in both HM mode AND LM mode |
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20 | * scope> Check gun tilt and shift |
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21 | * scope> Check rotation center and beam tilt pivot points in the HM mode |
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22 | * scope> Save the goniomenter position for the square with intact supporting film where the alignment was done and do the same for a square where the supporting film is broken (where bright/dark images can be acquired). |
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23 | 1 | Amber Herold | |
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25 | 11 | Amber Herold | 2. Start python-based leginon main program amd leginon |
26 | 1 | Amber Herold | client. See Chapter on [[Microscope Set-up|Start Leginon]]. |
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28 | 10 | Amber Herold | * scope> Set up [[MSI set-up in more details#Starting-Up|CCD camera]] |
29 | 11 | Amber Herold | * [[Run Leginon Client on the Microscope Computer|scope> Start Leginon Client]] |
30 | 10 | Amber Herold | * scope> If the following error message appears, DO NOT touch it, or Leginon Client will need restarting |
31 | 3 | Alex Kadokura | |
32 | 10 | Amber Herold | bq. _"Exception EAccessViolation in module adaExp.exe at...._ |
33 | _Access violation at address xxxx in module 'adaExp.exe'. Read of address xxxx"_ |
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34 | 1 | Amber Herold | |
35 | 10 | Amber Herold | * [[Start Leginon at your main working station remotely|main> start-leginon.py]] |
36 | * Leginon Setup Wizard> Create new or Select old session, connect to the instrument host, and start. |
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37 | 1 | Amber Herold | |
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39 | 11 | Amber Herold | 3. Start your preferred "MSI" application. |
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41 | 13 | Amber Herold | * Leginon/Application/Run> Select the "MSI" application you would like to use (Edge,T,Raster, or Tomography), the scope, and the main launcher. Click Run. |
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43 | h2. Import existing presets |
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45 | 13 | Amber Herold | * Leginon/Node Selector> Select "Presets Manager" node. |
46 | * Leginon/Presets Manager/Preset Selector> Import presets from a previous working session !http://emg.nysbc.org/software/leginon/images/icons/import.png! |
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47 | ** Select the TEM and camera to "Find" sessions closest to the condition of your data collection. |
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48 | ** Use shift-key to select a series of presets if needed. Hold-down to ctrl-key allows individual selection/deselection. |
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49 | ** Click "Import" button to import, the presets should appear in the Presets Manager/Preset Selector window. |
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50 | ** Click "OK" to exit "Import Presets" window. |
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51 | Note: Creating new preset and adjusting the preset are covered in Calibration Chapter under [[Creating Presets for the first time|Creating presets the first time]]. |
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52 | * The cycle order should match the sequence of data acquisition to minimize hysteresis. If the presets are imported from previous session, the cycle order is imported automatically. As an example, the cycle for naming scheme used here at NRAMM is gr -> sq -> hl -> fa -> fc -> en -> ef. |
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53 | 1 | Amber Herold | * Leginon/Presets Manager> Select a preset in the list box and then click "To Scope" !http://emg.nysbc.org/software/leginon/images/icons/instrumentset.png! to apply the settings to the scope and synchronize Leginon and the scope settings. |
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56 | 26 | Anchi Cheng | h2. Save the alignment and empty position in Leginon |
57 | 1 | Amber Herold | |
58 | 14 | Amber Herold | * scope> Recall the stored intact or empty position or go to such |
59 | 1 | Amber Herold | location. |
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61 | 14 | Amber Herold | * Leginon/Navigation/Toolbar/Stage Locations> !http://emg.nysbc.org/software/leginon/images/icons/stagelocations.png! Enter a New name to store Stage Location such as, for example, "empty" and "align", respectively. Save the xy coordinate only. To recall the position, select the named location under the same tool and click on "send to scope". |
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64 | h2. Grid preset image shift refinement |
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67 | 16 | Amber Herold | 1. Leginon/Presets Manager> Send a preset at an intermediate mag at HM mode as the reference (e.g. hl) "To Scope" !http://emg.nysbc.org/software/leginon/images/icons/instrumentset.png! |
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69 | 15 | Amber Herold | Note: sq preset in LM mode can be used if the image alignment between LM and HM |
70 | 14 | Amber Herold | is known to be good. |
71 | 1 | Amber Herold | |
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73 | 14 | Amber Herold | 2. scope> Move the grid to a position on the "align" square, either on a recognizable object or burn a good hole through ice at the center of the CCD as preparation for lower mag preset image shift correction. |
74 | 1 | Amber Herold | |
75 | 14 | Amber Herold | |
76 | 16 | Amber Herold | 3. Leginon/Presets Manager> Click on the tool "Align presets to each other" !http://emg.nysbc.org/software/leginon/images/icons/alignpresets.png!. |
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79 | 14 | Amber Herold | 4. Check, adjust, and save the preset(s) to be aligned. |
80 | 1 | Amber Herold | |
81 | 18 | Amber Herold | The left panel displays and navigate by "Stage Position" of the reference preset while the right panel displays and navigate by "Image Shift" of the preset to be aligned |
82 | 14 | Amber Herold | * Leginon/Presets Manager/Align Presets> Choose the reference (hl) preset on the left panel. |
83 | 1 | Amber Herold | |
84 | 14 | Amber Herold | * Leginon/Presets Manager/Align Presets> Choose the aligning (gr) preset on the right panel. |
85 | 1 | Amber Herold | |
86 | 14 | Amber Herold | * Leginon/Presets Manager/Align Presets> Select "Similar look across mags" mode at the top left corner of the window. This will use a smaller part of the CCD to acquire the image for the lower mag preset so that it appears to be magnified in the 512x512 image panel. |
87 | 1 | Amber Herold | |
88 | 14 | Amber Herold | * Leginon/Presets Manager/Align Presets> "Start" image acquisition. |
89 | 1 | Amber Herold | |
90 | 14 | Amber Herold | * Leginon/Presets Manager/Align Presets> If the recognizable object can not be found in both panels, go back to step one to choose a lower mag preset or a better object. |
91 | 1 | Amber Herold | |
92 | 14 | Amber Herold | * Leginon/Presets Manager/Align Presets> If the recognizable object can be found in both panels but not at the center on the reference left panel, navigate* it to the center using its "Click Tool". Wait for the move to complete and both images refreshed. |
93 | 1 | Amber Herold | |
94 | 14 | Amber Herold | * Leginon/Presets Manager/Align Presets> Navigate* the object on the right panel to the center using its "Click Tool".The new image shift is automatically saved at each move. |
95 | 1 | Amber Herold | |
96 | 14 | Amber Herold | * Repeat 7 if necessary. |
97 | 1 | Amber Herold | |
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99 | 14 | Amber Herold | Note: "Navigate" means selecting the navigate tool !http://emg.nysbc.org/software/leginon/images/icons/arrow.png! on the top right of image display and then double clicking the location of the new center that is translated by the given TEM Parameter. |
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103 | 14 | Amber Herold | h2. Grid preset stage position refinement |
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105 | 3 | Alex Kadokura | |
106 | 1 | Amber Herold | |
107 | 14 | Amber Herold | Stage position model Scale and Rotation is stable as long as the LM eucentric height defocus is set to zero before each session, although variation from day to day is normal. Check the stage movement calibration of gr preset by navigating it in Navigation node. Within 10% error is satisfactory since reaching a square does not require high accuracy. If not, perform stage model position _[[Modeled Stage Position calibration#Magnification-and-image-rotation-adjustment-of-the-model|scale and rotation calibration]]> at gr preset_ using "Calibration" application as following: |
108 | 1 | Amber Herold | |
109 | 14 | Amber Herold | _[[Modeled Stage Position calibration|Why goniometer model is better]]._ |
110 | 1 | Amber Herold | |
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113 | 14 | Amber Herold | 1. scope> Remove the objective aperture. |
114 | 1 | Amber Herold | |
115 | 14 | Amber Herold | 2. Leginon> Run "Calibrations" Application |
116 | 1 | Amber Herold | |
117 | 14 | Amber Herold | 3. Leginon/Presets Manager> Send "gr" preset to scope. |
118 | 1 | Amber Herold | |
119 | 14 | Amber Herold | 4. Leginon/GonioModeling/Settings/Measurement> Set axis=x, Points=5,Tolerance=25%, and Interval=1e-05 m. Default model label is the session name. |
120 | 1 | Amber Herold | |
121 | 14 | Amber Herold | 5. Leginon/GonioModeling/Settings/Modeling> Set "Scale and Rotation Adjustment Only"=Yes. Other settings will be set automatically once measurements are finished. |
122 | 1 | Amber Herold | |
123 | 14 | Amber Herold | 6. Leginon/GonioModeling> Click on !http://emg.nysbc.org/software/leginon/images/icons/cam_ruler.png! will start "Measure" the shifts of between points that are moved on x axis of the goniometer. |
124 | 1 | Amber Herold | |
125 | 14 | Amber Herold | 7. Leginon/GonioModeling> Click on !http://emg.nysbc.org/software/leginon/images/icons/play.png! will "Calibrate" the scale and rotation of the model on x axis of the goniometer. |
126 | 1 | Amber Herold | |
127 | 14 | Amber Herold | 8. Leginon/GonioModeling> repeat 4-7 with the axis set to y. |
128 | 1 | Amber Herold | |
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132 | h2. Acquire mosaic atlas of the grid |
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135 | 14 | Amber Herold | 1. scope> Remove the objective apperture. |
136 | 1 | Amber Herold | |
137 | 14 | Amber Herold | 2. Leginon/Grid Targeting> Click "Settings" to check the preset used, the size and name of the atlas. |
138 | 1 | Amber Herold | |
139 | 14 | Amber Herold | 3. Leginon/Grid Targeting> Click "Calculate Atlas" !http://emg.nysbc.org/software/leginon/images/icons/calculate.png! |
140 | 1 | Amber Herold | |
141 | 14 | Amber Herold | 4. Leginon/Grid Targeting> Click "Submit Targets" !http://emg.nysbc.org/software/leginon/images/icons/play.png! to begin acquiring the atlas. |
142 | 1 | Amber Herold | |
143 | 14 | Amber Herold | 5. Leginon/Square Targeting> Follow the atlas progress in the display. (***There is no sign at the end currently other than that nothing happens for a long time and that the atlas dimension is the same in other direction when it is completed.) |
144 | 1 | Amber Herold | |
145 | 14 | Amber Herold | 6. scope> Insert the objective aperture. |
146 | 1 | Amber Herold | |
147 | 14 | Amber Herold | 7. Leginon/Presets Manager/Send an HM preset "To Scope" several times to cycle all magnifications and warm up the objective lens to minimize the image shift hysteresis in switching between LM and HM modes. If the scope has been in LM mode for a long time, wait for at least 30 min before final image shift refinement of needed LM preset during acquisition (i.e. sq) |
148 | 1 | Amber Herold | |
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152 | 14 | Amber Herold | h2. Determine the proper exposure condition for exposure presets |
153 | 1 | Amber Herold | |
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156 | 17 | Amber Herold | Exposure presets are the base presets for data collection. To keep it to low dose condition such as 10 electron/Angstrum, measurement of dose is performed and the exposure time is adjusted. If scope or camera parameters are changed during the session, the dose value is no longer valid and should be remeasured. |
157 | 1 | Amber Herold | |
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159 | 14 | Amber Herold | 1. Leginon/Navigation/Toolbar/Stage Locations !http://emg.nysbc.org/software/leginon/images/icons/stagelocations.png! Send the "empty" position "To scope". |
160 | 1 | Amber Herold | |
161 | 14 | Amber Herold | 2. Leginon/Presets Manager/Preset Selector> Select a high mag exposure (i.e. "en" or "ef"), confirm that the parameters are what you desire for the experiment, and then send "To Scope" !http://emg.nysbc.org/software/leginon/images/icons/instrumentset.png!. |
162 | 3 | Alex Kadokura | |
163 | 14 | Amber Herold | 3. scope> Expand the beam to at least cover the CCD (> medium circle mark on the viewing screen) and to the size you desire. |
164 | 1 | Amber Herold | |
165 | 14 | Amber Herold | 4. Leginon/Presets Manager/Preset Selector> If you have changed the size or location of the beam or other preset scope parameters at the scope, you must retrieve |
166 | the preset parameter "From scope" !http://emg.nysbc.org/software/leginon/images/icons/instrumentget.png! first before checking and saving the dose value. |
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167 | 1 | Amber Herold | |
168 | 14 | Amber Herold | 5. Leginon/Presets Manager/Preset Selector> "Acquire dose image" !http://emg.nysbc.org/software/leginon/images/icons/acquire.png! for the selected preset. |
169 | 1 | Amber Herold | |
170 | 14 | Amber Herold | 6. Leginon/Presets Manager/Dose Image> Check the "Dose" value. |
171 | 1 | Amber Herold | |
172 | 14 | Amber Herold | 7. Leginon/Presets Manager/Dose Image> If desired dose is known, fill in the value and then click "Match". This will change exposure time to give a close match of |
173 | 1 | Amber Herold | the value entered. |
174 | 3 | Alex Kadokura | |
175 | 14 | Amber Herold | 8. Leginon/Presets Manager/Dose Image> If not satisfied with the value, click "No", change the exposure time by editing the preset, and try again (No need to send |
176 | 1 | Amber Herold | the preset "To scope" again for the repeat) |
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178 | 14 | Amber Herold | 9. If the scope can not give such illumination to satisfy dose, exposure time and area requirement, alignment is required or change of gun lens or spot size is needed. |
179 | 1 | Amber Herold | |
180 | 14 | Amber Herold | 10. Leginon/Presets Manager> Copy the preset parameter to the other exposure preset. |
181 | 1 | Amber Herold | |
182 | 14 | Amber Herold | * Preset Selector> send the preset that has the dose measurement "To Scope" !http://emg.nysbc.org/software/leginon/images/icons/instrumentset.png! |
183 | * Preset Selector> select the preset to receive the copy |
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184 | * Preset Selector> retrieve the parameters "From Scope" !http://emg.nysbc.org/software/leginon/images/icons/instrumentget.png! for the selected preset. |
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185 | * Preset Selector/Edit> !http://emg.nysbc.org/software/leginon/images/icons/edit.png! Modify defocus for the preset after the retrieval. |
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186 | 1 | Amber Herold | |
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189 | 14 | Amber Herold | h2. Update bright/dark correction images for exposure presets |
190 | 1 | Amber Herold | |
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193 | 14 | Amber Herold | Exposure presets are the base presets for data collection. To make sure the best correction is applied to them, the correction images should be acquired prior to the experiment at session. Since no correlation is normally done on these images, only one [[Bright and Dark reference images#Correction-Channels|channel]] is needed. Normalization image might need to be [[Bright and Dark reference images#Correction-Planmodified]] if a large chunk of region is blocked by a fallen debris. |
194 | 1 | Amber Herold | |
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196 | 14 | Amber Herold | 1. Leginon/Presets Manager> Select a dose measured high mag exposure (i.e. "en" or "ef"), and then send "To Scope". |
197 | 1 | Amber Herold | |
198 | 14 | Amber Herold | 2. Leginon/Correction/Settings> |
199 | 1 | Amber Herold | |
200 | 14 | Amber Herold | * Select TEM and camera to match that will be used. |
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202 | * Camera Configuration: The exposure preset camera configuration |
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203 | 1 | Amber Herold | |
204 | 14 | Amber Herold | 3. Leginon/Correction/Settings> Click OK to exit settings. |
205 | 1 | Amber Herold | |
206 | 14 | Amber Herold | 4. Leginon/Correction/Toolbar> Select "Raw image " from the pull down list of acquisition modes and then click on "Acquire" button next to the selector to view an image that is not corrected. |
207 | 1 | Amber Herold | |
208 | 14 | Amber Herold | 5. Leginon/Correction/Toolbar> Select "Dark reference" in the acquisition mode selector and then click "Acquire" to acquire the Dark reference image for this particular camera configuration. |
209 | 1 | Amber Herold | |
210 | 14 | Amber Herold | 6. Leginon/Correction/Toolbar> Select "Bright reference" and repeat the acquisition to obtain the Bright reference. |
211 | 1 | Amber Herold | |
212 | 14 | Amber Herold | 7. Leginon/Correction/Toolbar> Select "Corrected image" and then "Acquire" to view the corrected image. A corrected image should be free of artifacts and have |
213 | 1 | Amber Herold | smaller standard deviation than the raw image, in general. |
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215 | 14 | Amber Herold | 8. Leginon/Correction/Toolbar> If there are regions consistently over normalizes the acquired image, either select the region and use the "+" tool to "Add Region to Bad Pixel List", or click on !http://emg.nysbc.org/software/leginon/images/icons/stagelocations.png! to "Add Extreme Points to Bad Pixel List" one of more times to find the outliers. Right click on a shown bad pixel removes it from the list. |
216 | 1 | Amber Herold | |
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219 | h2. Medium mag preset image and beam shift refinement |
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222 | 14 | Amber Herold | Preset image shift alignment can be done follow the same procedure as the section "[[MSI set-up in more details#Grid-preset-image-shift-refinement|Grid preset image shift refinement]]" but is more efficient using the automated mode as follows. |
223 | 1 | Amber Herold | |
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225 | 22 | Amber Herold | 1. Leginon/Presets Manager> Send one of the exposure preset as the reference (e.g. en) "To Scope" !http://emg.nysbc.org/software/leginon/images/icons/instrumentset.png! |
226 | 1 | Amber Herold | |
227 | 14 | Amber Herold | 2. scope> Move the grid to a position on the "align" square, either on a recognizable object or burn a good hole through ice at the center of the CCD as preparation for lower mag preset image shift correction. |
228 | 1 | Amber Herold | |
229 | 14 | Amber Herold | 3. Leginon/Presets Manager> Click on the tool "Align presets to each other" !http://emg.nysbc.org/software/leginon/images/icons/alignpresets.png!. The tool automatically select the reference preset that was selected in the Presets Manager mainpanel and choose a preset at closest magnification as the starting aligning preset. |
230 | 1 | Amber Herold | |
231 | 14 | Amber Herold | 4. Leginon/Presets Manager/Align Presets> Click "Start" to acquire the first image pair. |
232 | 1 | Amber Herold | |
233 | 14 | Amber Herold | 5. Check, adjust, and save the preset(s) to be aligned. |
234 | 1 | Amber Herold | |
235 | 19 | Amber Herold | The left panel displays and navigate by "Stage Position" of the reference preset while the right panel displays and navigate by "Image Shift" of the preset to be aligned |
236 | 1 | Amber Herold | |
237 | 14 | Amber Herold | * Leginon/Presets Manager/Align Presets> Changing Selection of "Similar look across mags" or "Full CCD" mode at the top left corner of the window will cause reacquisition of the image pair in the automated mode. |
238 | 1 | Amber Herold | |
239 | 14 | Amber Herold | * Leginon/Presets Manager/Align Presets> If the recognizable object can be found in both panels but not at the center on the reference left panel, navigate* it to the center using its "Click Tool". Wait for the move to complete and both images refreshed. |
240 | 1 | Amber Herold | |
241 | 14 | Amber Herold | * Leginon/Presets Manager/Align Presets> Navigate* the object on the right panel to the center using its "Click Tool". The new image shift is automatically saved after each move. |
242 | 1 | Amber Herold | |
243 | 14 | Amber Herold | * Repeat 3 if necessary. |
244 | 1 | Amber Herold | |
245 | 14 | Amber Herold | * Leginon/Presets Manager/Align Presets> Click "Continue" to save the alignment to all presets of the same magnification and continue to the next preset pair. |
246 | 1 | Amber Herold | |
247 | 14 | Amber Herold | 6. Repeat the process if desired. |
248 | 1 | Amber Herold | |
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251 | 8 | Anchi Cheng | h2. Go to MSI Operation section |
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253 | 1 | Amber Herold | |
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255 | 21 | Amber Herold | The setup that requires you to be next to the microscope is completed. Go to the section on [[MSI Operation|MSI Operation]] You can come back and continue on this section after you have learned how to go through the operation without automated hole finding. |
256 | 1 | Amber Herold | |
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259 | h2. Hole Targeting Set-up |
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262 | 21 | Amber Herold | We can use any image that is already loaded into Hole Targeting to adjust these parameters. The values in this node vary so much from grid to grid that the best way to find these parameters is by going through a step-by-step trial-and-error process. To see the final acquisition and focus targets, enable only the Original, acquisition, and focus display selections. Refer to Introduction Chapter on [[MSI set-up in more details|how the various tools behave in the control panel]]. |
263 | 1 | Amber Herold | |
264 | 4 | Alex Kadokura | |
265 | 21 | Amber Herold | The loaded image normally comes from the image acquired through "Square" node when you submit targets on the grid atlas in "Square Targeting" node. You can also do a practice run using the default image comes with Leginon. Leave the filename entry blank when you click "Load" in the hole finding settings window for the step "Original". The default values in the node gives an example of reasonable target finding with such default image. |
266 | 1 | Amber Herold | |
267 | |||
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269 | Setting up Hole Targeting Depends on which MSI flavor you are running: |
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272 | 21 | Amber Herold | 1. [[Hole Targeting Set-up for MSI-Edge|MSI-Edge]] uses template matching of the edge image |
273 | 1 | Amber Herold | |
274 | 21 | Amber Herold | 2. [[JAHC finder nodes set-up|MSI-T]] uses template matching of the original image |
275 | 1 | Amber Herold | |
276 | 21 | Amber Herold | 3. [[raster finder nodes set-up|MSI-Raster]] creates raster points from a given origin |
277 | 1 | Amber Herold | |
278 | 21 | Amber Herold | 4. [[Initial MSI application preference setup#Settings-that-are-the-same-as-in-other-MSI-applications-in-queuing-mode|MSI-Tomography]] does not use automatic target finding. Therefore, it does not requires set up. |
279 | 1 | Amber Herold | |
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282 | h2. Exposure Targeting Set-up |
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286 | 21 | Amber Herold | Like hole targeting, we can use any image that is already loaded into Exposure Targeting to adjust these parameters. |
287 | 1 | Amber Herold | |
288 | Setting up Exposure Targeting Depends on which MSI flavor you are running. |
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291 | 21 | Amber Herold | 1. [[Hole Targeting Set-up for MSI-Edge|MSI-Edge]] uses template matching of the edge image |
292 | 1 | Amber Herold | |
293 | |||
294 | 21 | Amber Herold | 2. [[JAHC finder nodes set-up|MSI-T]] uses template matching of the original image |
295 | 1 | Amber Herold | |
296 | 4 | Alex Kadokura | |
297 | 21 | Amber Herold | 3. [[raster finder nodes set-up|MSI-Raster]] creates raster points from a given origin |
298 | 1 | Amber Herold | |
299 | |||
300 | 21 | Amber Herold | 4. [[Initial MSI application preference setup#Settings-that-are-the-same-as-in-other-MSI-applications-in-queuing-mode|MSI-Tomography]] does not use automatic target finding. Therefore, it does not requires set up. |
301 | 1 | Amber Herold | |
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304 | h2. Autofocus Test |
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307 | 4 | Alex Kadokura | |
308 | 21 | Amber Herold | There are two focusers in MSI. "Z Focus" node usually moves stage in Z direction to Eucentric height at lower mags. "Focus" node usually just correct the defocus in the way equivalent to turning the focus adjustment knob on the scope at a higher mag. The defocus measurements in both nodes are based on the same principle of beam-tilt induced image shift. A correlation (normally phase) is used to obtain the image shift. Therefore, it is important to test and preform autofocus measurement at area with recognizable feature that does not move significantly during the defocus measurement. |
309 | 1 | Amber Herold | |
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311 | 21 | Amber Herold | The test use "Manual_after" focus step in the focus sequence for checking. Two procedures can be used to reach a area for good correlation measurements: |
312 | 1 | Amber Herold | |
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314 | 21 | Amber Herold | 1. Using Simulate Target tool |
315 | 1 | Amber Herold | |
316 | 21 | Amber Herold | * Leginon/Presets Manager> Send the autofocus preset "To Scope" |
317 | 1 | Amber Herold | |
318 | 21 | Amber Herold | * scope> Move to an area that has recognizable feature on the carbon support at the center. |
319 | 1 | Amber Herold | |
320 | 21 | Amber Herold | * Leginon/(Z) Focus/Focus Sequence !http://emg.nysbc.org/software/leginon/images/icons/focus_sequence.png! > Enable desired autofocus and "Manual_after" focus steps. |
321 | 1 | Amber Herold | |
322 | 21 | Amber Herold | * Leginon/(Z) Focus/Toolbar> Click Simulate Target !http://emg.nysbc.org/software/leginon/images/icons/simulatetarget.png! and follow the autofocus |
323 | 1 | Amber Herold | |
324 | |||
325 | 21 | Amber Herold | 2. Submitting focus target from Hole Targeting |
326 | 1 | Amber Herold | |
327 | 21 | Amber Herold | Note: To use this procedure, there has to be a grid mosaic, and the Hole Targeting should allow user verification of targets |
328 | 1 | Amber Herold | |
329 | |||
330 | 23 | Amber Herold | * Leginon/(Z) Focus/Focus Sequence !http://emg.nysbc.org/software/leginon/images/icons/focus_sequence.png! > Enable desired autofocus and "Manual_after" focus steps. |
331 | 1 | Amber Herold | |
332 | 21 | Amber Herold | * Leginon/Square Targeting> Select a test square and submit it !http://emg.nysbc.org/software/leginon/images/icons/play.png! |
333 | 1 | Amber Herold | |
334 | 21 | Amber Herold | * Leginon/Hole Targeting> Submit a focus target to test autofocus in Z Focus node and an acquisition target for the next stage. Both targets should be in an area that has recognizable feature with carbon support. |
335 | 1 | Amber Herold | |
336 | 21 | Amber Herold | * Leginon/Z focus> follow the autofocus test in this node. |
337 | 1 | Amber Herold | |
338 | 21 | Amber Herold | * Leginon/Exposure Targeting> Submit a focus target at a location that has recognizable feature with carbon support to test autofocus in Focus node. |
339 | 1 | Amber Herold | |
340 | 21 | Amber Herold | * Leginon/Focus> follow autofocus test in this node. |
341 | 1 | Amber Herold | |
342 | |||
343 | 21 | Amber Herold | Check the performance of autofocus by the following criteria: |
344 | 1 | Amber Herold | |
345 | |||
346 | 21 | Amber Herold | 1. During autofocusing: |
347 | 1 | Amber Herold | |
348 | 21 | Amber Herold | * Display the image and correlation peak: The observed movement of images upon beam tilt should be consistent with the correlation peak and the edge of the beam should not be visible in any of the beam-tilt images. |
349 | 1 | Amber Herold | |
350 | 21 | Amber Herold | * Display the correlation image: The correlation peak at its correct position should be significantly higher than background. |
351 | 1 | Amber Herold | |
352 | |||
353 | 21 | Amber Herold | 2. During manual focusing |
354 | 1 | Amber Herold | |
355 | 21 | Amber Herold | Manual Focusing can be activated either by enabling "Manual_before/Manual_after" steps or directly when leginon is idle using (Z) Focus/Toolbar/MF. To activate manual focus directly, it is necessary to send to scope the preset of the choice (normally fc) first, though. |
356 | 1 | Amber Herold | |
357 | 21 | Amber Herold | * MF> The FFT should show visible and isotropic Thon ring according to the fc preset defocus. |
358 | 1 | Amber Herold | |
359 | 21 | Amber Herold | * MF/Toolbar> Make sure the move type is "Defocus" and the set value in the box is 0 m then click "Set instrument" botton !http://emg.nysbc.org/software/leginon/images/icons/instrumentset.png!. This will set the defocus to the current 0. |
360 | 1 | Amber Herold | |
361 | 21 | Amber Herold | * MF> The perfect autofocus result is that the FFT indicates that the scope is at Gaussian focus. |
362 | 1 | Amber Herold | |
363 | 21 | Amber Herold | * For relative movement, press "+" or "-". The step size is determined in MF/Toolbar/Settings> |
364 | 2 | Amber Herold | |
365 | 25 | Amber Herold | * To reset defocus to zero at the new focus, press the so-named button !http://emg.nysbc.org/software/leginon/images/icons/instrumentsetnew.png!. |
366 | 2 | Amber Herold | |
367 | 21 | Amber Herold | * Clicking the Stop button !http://emg.nysbc.org/software/leginon/images/icons/stop.png! exits the manual check. |
368 | 2 | Amber Herold | |
369 | |||
370 | |||
371 | 1 | Amber Herold | ______ |
372 | |||
373 | [[MSI Quick-start|< MSI Quick-start]] | [[Special Operation setup|Special Operation setup >]] |
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374 | |||
375 | ______ |