Tasks menu commands

The Tasks menu offers the following commands:

A Note on Tasks in Low Dose Mode

Tasks that use lower magnification tracking will generally use View images in Low Dose mode, although some have menu entries to control which area is used.  However, each such task has a required minimum field of view when run outside of Low Dose, which sets a minimum magnification at which it should be run.  When the View magnification is higher than that minimum, and the Search magnification is lower than View and not in LM mode, the program will use Search instead to minimize the risk of excessive shifts between images.

Rough Eucentricity command (Tasks - Eucentricity submenu)

This command can be used to adjust the Z height to the eucentric point when the Z height might be off by more than 5-10 microns, which can often be the case when the specimen rod has just been inserted. This procedure will find eucentricity to within about 0.5 micron. This is good enough for tuning the microscope but not as accurate as the adjustment provided by the Eucentric - Fine command, which should be run before starting a tilt series.

This procedure does not make any attempt to keep the starting specimen location in the field of view.

The rough eucentricity procedure first switches to a low magnification (or, in Low Dose mode, it uses the parameters of the View or Search area, as described above). It takes pictures to determine how far the specimen moves laterally upon tilting, starting with a very small tilt increment and working up to a total increment of 8 degrees. It may adjust the Z height more than once before finishing.

Fine Eucentricity command (Tasks - Eucentricity submenu)

This command can be used to adjust the Z height to the eucentric point when it is already within 5-10 microns. It is thus a suitable procedure to use if you have moved to a new location on the specimen grid since the last time eucentricity was adjusted. It is recommended that you run this command before starting a tilt series.

This procedure does not make any attempt to keep the starting specimen location in the field of view. It can be run when not at zero tilt, and the program will return the stage to your pre-existing tilt at the end.

The fine eucentricity procedure first switches to a low magnification (or, in Low Dose mode, it uses the parameters of the View area or Search area, as described above). It then takes pictures at a set of tilts between -24 and +24 degrees. From the image displacements during this series, it can compute both the distance in Z from the eucentric point and the lateral displacement of the tilt axis from the optical axis. If image displacements become too large, the program will adjust Z height then repeat the procedure from the beginning.

For specialized uses it may be desirable to change the maximum angle or tilt increment.  These parameters can be changed with the properties 'EucentricityMaxFineAngle' and 'EucentricityFineIntervalAtZero'.

Both Rough and Fine command (Tasks - Eucentricity submenu)

Use this command to run the rough eucentricity procedure followed immediately by the fine eucentricity procedure. This sequence not does make any attempt to keep the starting specimen location in the field of view.

Refine & Realign command (Tasks - Eucentricity submenu)

Use this command to run the fine eucentricity procedure while retaining an item of interest centered in the field of view. The program will first take a low-magnification reference picture, then do the fine eucentricity procedure, then take another picture at the original tilt angle and use that to restore the alignment. In Low Dose mode, the reference picture will be taken of the View or Search area, as described above. It is not necessary to be at zero tilt to run this command.

Set Tilt Axis Offset command (Tasks - Eucentricity submenu)

Use this command to set the offset of the tilt axis from the optical axis, which is used to keep image shift centered on the optical axis. If the tilt axis is more than a micron from the optic axis, it is desirable to set this offset to minimize specimen movements in Y and Z during tilting. The offset should be set as a system-wide property. However, if this property setting is not adequate, or if the offset varies significantly from place to place, this command can be used to set the offset. It is placed in this menu because the lateral offset from the Refine Eucentricity procedure indicates how much the offset should be changed. When the command is selected, a dialog box comes up informing you of the current tilt axis offset, and the total offset implied by the lateral displacement reported in the last run of Refine Eucentricity, if any. The default value in the box is the implied total offset (or just the current offset, if Refine Eucentricity has not been run.) A tilt axis offset has an effect only if you have selected 'Center image shift on tilt axis' in the Image Alignment and Focus control panel.

Set Axis Offset Automatically from Fine command (Tasks- Eucentricity submenu)

This command toggles an option to have the tilt axis offset set automatically to the total lateral offset determined from running the fine eucentricity procedure.  It will use the same number that appears in the entry box when using the preceding command.

Eucentricity by Focus command (Tasks - Eucentricity submenu)

Use this command to set the Z height to the eucentric point with a procedure that sets an absolute focus value then measures the defocus and adjusts Z until the measured defocus matches the appropriate value.  Both the absolute focus and the measured defocus are recorded with a calibration procedure run from the Eucentricity by Focus Setup dialog opened by the next menu item.  Defocus can be measured with either Focus or View images in Low Dose mode, with a desired amount of defocus offset.

Setup Eucentricity by Focus command (Tasks - Eucentricity submenu)

Use this command to open the Eucentricity by Focus Setup dialog and calibrate the focus targets for the Eucentricity by Focus command as well as set a few parameters controlling how it operates.

Set Low Dose Areas command (Tasks menu)

Use this command to open the  Low Dose Area Options for Tasks dialog and select which areas are used for various tasks in Low Dose mode, including Rough and Fine Eucentricity, Reset IS & Realign, and Walkup.

Set Intensity command (Tasks menu)

Use this command to change the beam intensity so as to achieve a desired mean level in images taken with Record parameters. When you select this command, the program first computes the zero-tilt mean (mean of the tilt-foreshortened area) of the image in buffer A. It then takes account of the exposure time and binning of that image to determine what the zero-tilt mean of a Record image would have been at the current beam intensity. The program presents a dialog box in which you can enter the new intensity; the default value in this box is the computed mean for a Record image at the current intensity. This value will give you some idea of how big a change you are requesting. If you accept this default, nothing will change. Enter the desired mean level.

Since the program changes intensity by condensing the beam, it is possible that the beam will become too small to cover the camera area. If you increase the intensity by a substantial amount, you should check whether the beam still covers the area before going on to tasks that rely on full exposures.

In Low Dose mode, this command will change the intensity setting for the Low Dose area from which the image was acquired, regardless of whether 'Continuous update' is on in the Low Dose control panel.

Set Dose Rate command (Tasks menu)

Use this command to change the beam intensity to give a desired dose rate on the camera, in electrons per physical pixel per second, based on the counts in the image in buffer A.  The mean of the tilt-foreshortened area will be used, as for the Set Intensity command.  The programs presents a dialog box showing the dose rate in that image and asking for the new value.  In Low Dose mode, this command will also change the intensity setting for the Low Dose area from which the image was acquired.

Electron Dose command (Tasks menu)

Use this command to calibrate electron dose for a particular spot size. This calibration is also specific to the probe mode for Thermo/FEI scopes. It is automatically saved in a special short-term calibration file and is only good for a limited period of time (1 day by default, but this time can be set with the property 'DoseLifetimeInHours'). The calibration requires that beam intensity be calibrated and that the SerialEMproperties.txt file contain an entry for CountsPerElectron for the camera.

Before activating the command, take an image of the blank beam with a reliable exposure time (e.g., 0.2 or more seconds) and with the beam in the calibrated range for the current spot size.  Specifically, the intensity has to be within the directly calibrated range, not in the range where the calibration can be extrapolated, so the 'SPOT' label in the microscope status panel should be darker blue, not light cyan. Select the command and press OK. The program will report the dose rate (electrons per square Angstroms per second) of the beam intensity at which that image was taken.  Or, if intensity was not in the directly calibrated range, there will be a message telling you what that range is.

The dose calibration will be most accurate if the spot size is normalized (on a Thermo/FEI scope). To do this, open the Normalizations panel in the Microscope User Interface, select TEM Spotsize, and check Spotsize. Then change spot size to get a normalized beam, take a picture, and calibrate the dose with it. By the same token, the actual measurement of dose at a particular beam setting will be more accurate if the spot size is normalized whenever you change spot sizes. Hysteresis in the second condenser lens will also reduce the accuracy of a dose measurement, but this is more difficult to control for.

The dose calibration is specific to the side of beam crossover on which the calibration image was taken. Once a calibration is present, the program can compute the dose for any image acquisition when beam intensity is within the calibrated range, taking into account drift settling time as well as CCD exposure time. If relative spot intensities have been calibrated, then a calibration at a single spot size will allow dose to be computed at any spot size. Note that dose is automatically calibrated when you take a gain reference, unless you deselect this option in the Gain Reference dialog .

When there is an electron dose calibration, the estimated dose at the specimen will be displayed in several places:

1. The dose for a low dose acquisition area will appear in the upper right of the Low Dose control panel, with a continuous readout if 'Continuous update' is selected.
2. The dose for the current camera exposure time, in the lower right of the Camera Setup dialog .
3. When a direct electron detector is the selected camera, the dose rate at the specimen, per unbinned pixel per second, will appear in the Options portion of the Microscope Status panel.

Purge Old Calibs command (Tasks menu)

This command will remove any dose calibrations done before the current instance of the program was started. You would use this if you do a dose calibration (or take a gain reference) and then want to remove any earlier calibrations. Dose calibrations older than 24 hours are ignored when the program starts. Also, when dose is calibrated, other calibrations older than 12 hours are then removed.

Set Aperture Size command (Tasks menu)

Use this command to inform the program of the current size of the C2 aperture on a Titan microscope, where the illuminated area read from the microscope and used for intensity is scaled by this aperture size when the aperture is changed.  The command is disabled if the program determines on startup that it can control the apertures through UTAPI scripting, or if the property 'MonitorC2ApertureSize' is set to 2 and aperture control is available otherwise.  The program will scale the intensity values in the beam intensity, spot intensity, and crossover calibrations so that they are valid for this aperture size.   Once the aperture size is set either with this command or after one of those calibrations, it will be remembered between sessions in the short term calibration file.  If the aperture is changed only through commands from SerialEM, this command should not be needed, because the program will scale the intensities after each C2 aperture change (unless 'MonitorC2ApertureSize' is set to 0 to disable this feature).

Move Beam command (Tasks menu)

Use this command to move the beam by a controlled amount. You can use it to center the beam by first taking a picture with the beam condensed enough to show much of its boundaries. Then click the left mouse button to set a display marker point in the center of the beam. Next, select this command, and the center of the beam will be moved to the center of the field. In general, the beam will be moved by the amount that would move the marker point to the center of the field.

After you use this command, you need to take another picture to see the position of the beam.

Center Beam command (Tasks menu)

You can use this command to center the beam if the edge of the beam is fairly sharp and appears in the currently active image. The program will analyze the image to find the beam edge then solve for a circle that fits the edge. It will then move the beam to move the center of that circle toward or to the center of the image. The solution will be most accurate if the beam edge appears in 3 or 4 corners of the image, and will be fairly accurate when there is beam edge in two corners of the image. With an edge in only one quadrant, the program will be increasingly conservative in how much it moves the beam, the smaller the piece of edge is. When the edge occupies less than 20 degrees, the beam will simply be moved enough to get the edge out of the field, so do not expect the beam to be centered in such situations. This command will not work if the beam edge is not relatively sharp, so it will not work with some non-FEG scopes.

If the log says 'No beam edges detectable in this image' for an image with clear edges, check the counts in the region otside the beam compared to inside the beam.  The program requires that the counts outside fall below 15% of the mean inside.  If the background counts outside the beam are too high, set the property 'FindBeamOutsideFrac' to an appropriate value above 0.15.

Autocenter Beam command (Tasks menu)

You can use this command to center the beam automatically by taking an image with the beam condensed so that its position can be detected reliably either from the edges of the beam or from the average location of the beam in the image. You must first set up the beam intensity at the current or a nearby magnification and spot size, using the Beam Autocenter Setup dialog box . The program will condense the beam, take one or two images depending on the centering method, and restore the beam.

Setup Autocenter command (Tasks menu)

Use this command to open the Beam Autocenter Setup dialog box and set the beam intensity that will be used for autocentering.

Walk Up command (Tasks menu)

Use this command to go from one tilt angle to another while retaining an item of interest centered in the field of view. Typically this would be used to go from zero tilt to a high tilt angle. After you select the command, you will encounter a dialog box in which to enter the destination tilt angle. This destination angle will be saved and will appear as the default angle the next time you invoke the command.

This procedure involves taking alignment pictures at a series of tilt angles, with a tilt interval that is quite coarse at low angles and finer at high angles. Image shift will be reset whenever it gets too high, but no autofocusing is done. The pictures are taken with the Trial parameter set and, if in Low Dose mode, the pictures are of the Trial area. If the magnification is too high for reliable tracking, the pictures will be taken at a lower magnification.

If you choose a destination angle that is in the opposite direction from the last direction of tilt, the routine will run the Reverse Tilt procedure first to avoid losing the area of interest.

Walk Up & Anchor command (Tasks menu)

Use this command to do the same operation as in the Walk Up command but also leave an image behind at a specified intermediate tilt angle. The image will be left in the last image buffer and can be used as an 'anchor' for alignment during a tilt series. The angle for the anchor image will be saved as the default for the next use of this command and is the same as the angle that appears for an anchor image in the Tilt Series Setup dialog box .

Use View in LD command (Tasks menu)

Use this command to select whether the Walk Up routine will be run with the View or Search area (as described above) instead of the Trial area when in Low Dose mode.

Set Increments command (Tasks menu)

Use this command to set the minimum and maximum tilt increments for the Walk Up operation. The maximum increment is used at zero tilt, and the increment at any tilt angle is either this maximum times the cosine of the angle, or the minimum increment, whichever is greater. In STEM mode, you can also set the tilt interval between autofocusing during a Walk Up.  Your settings for these values are retained between sessions of SerialEM.

Setup Cooker command (Tasks menu)

Use this command to open the Specimen Cooking Setup dialog box and set the magnification, spot size, and intensity to use for pre-exposing the specimen.

Cook Specimen command (Tasks menu)

Use this command to pre-expose the specimen for the desired electron dose or time, using the parameters set in the Specimen Cooking Setup dialog box . The program will lower the screen to expose the specimen, report in the log window as each tenth of the exposure is finished, and continually update the status bar with the expected time remaining.

Assess Angle Range command (Tasks menu)

Use this command to acquire a small series of images at high tilt in order to determine the limits of the tilt range for a tilt series. There are options that allow a series to be acquired as quickly as possible, or more slowly and carefully, depending on what you wish to see in the high tilt images. These options are set in the Tilt Series Range Finder dialog box.

Assess Interset Shifts command (Tasks menu)

In STEM mode, use this command to measure the shifts between Record and other images with current camera parameters.  These shifts are compensated during tilt series acquisition to make sure that tracking and focusing images are centered on the Record area.  The shifts are specific to the magnification and the image size and binning, and also depend on exposure time when the scan rate is relatively high.  The shifts are not saved for different conditions, so this command needs to be run as part of the routine tasks before starting a tilt series, after camera parameters have been set up.  The Record exposure time can be varied during the tilt series, but the other exposure times should be kept constant, or else the shifts will not be considered valid.

When not in Low Dose mode, the routine will measure the shift for a Trial image and for a Focus image relative to Record.  When in Low Dose mode, the routine will always measure a shift for a Preview image.  It will then measure a shift for a Trial if Trial area is centered on Record, and a shift for Focus if the Focus area is centered on Record.  At the end, you should scroll through the images to make sure they are well-enough aligned.  If the image in A is not aligned, you can align it with the right mouse button then use the Revise/Cancel Shifts command.  If a different image is not aligned, you can try the command again with a new image area, or eliminate the shifts with the Revise/Cancel Shifts command.

Revise/Cancel Shifts command (Tasks menu)

Use this command either to indicate that an interset shift should be taken from the alignment of the image in buffer A, or to eliminate the last measured interset shifts.  You will first be asked if you want the revise a shift based on the alignment of the image in A.  If you answer No, you will then be asked if you want to eliminate the interset shifts.

Reset IS & Realign command (Tasks menu)

Use this command to reset the microscope image shift (IS) while retaining an item of interest centered in the field of view. The program will take a reference picture, reset the image shift to zero and move the stage to compensate, then take another picture and align it to the reference picture. At the end, image shift will no longer be zero because of this alignment; in fact, it can be as much as one-quarter of the original image shift because of inaccuracy in stage movement. The reference picture will be taken at low magnification or, if in Low Dose mode, with the View or Search parameters, as described above.

If you just want to reset image shift without having a feature stay centered, use the Reset Shifts button in the Image Alignment & Focus panel .

Set Iteration Limit command (Tasks menu)

Use this command to change the criterion that the Reset IS & Realign procedure uses to decide whether to reset the shift and realign a second time. After resetting the shift, the program takes an image and aligns it to image taken before the reset. If image shift is still greater than this criterion, it will be reset again, and another image taken and aligned.

Setup VPP Conditioning command (Tasks menu)

Use this command to open the Phase Plate Conditioning Setup dialog box and set parameters for conditioning a Volta phase plate. The dialog allows one to start the conditioning operation, which can also be run with the ConditionPhasePlate script command.

Setup Wait for Drift command (Tasks menu)

Use this command to open the Wait for Drift Setup dialog box and set parameters for the Wait for Drift task, which can be run from tilt series and with the script command DriftWaitTask to measure drift and wait until it reaches a selected level.

Reverse Tilt command (Tasks menu)

Use this command to reverse the direction of tilting while retaining an item of interest centered in the field of view. For example, if you tilt up to a high angle, center your item of interest, then try to start a tilt series by just tilting back down, the specimen may move considerably because of backlash in the stage. This problem can be avoided by using this command. The program will take a reference picture, tilt far enough in the current direction to work out the backlash, tilt back to the starting angle, then take another picture and align it to the reference picture. The reference picture will be taken at low magnification or, if in Low Dose mode, with the View or Search parameters, as described above.

Setup Scope Management command (Tasks menu)

Use this command to open the Dewar and Vacuum Management dialog, which allows you to set parameters for a routine that can manage periodic operations on the microscope that might interfere with data collection, either vacuum pumps running on a Thermo/FEI scope, or nitrogen dewar filling on various scopes.  The routine can check whether some events have begun and wait until they are done, or start some events proactively and wait until completion.

Verbose command (Tasks menu)

Use this command to toggle diagnostic output to the Log window from the various procedures run from the Tasks menu.