Camera menu commands
The Camera menu offers the following commands:
|Parameters||Opens dialog box to set parameters for image acquisition.|
|Prepare Gain Ref||Opens dialog box to acquire a gain reference in SerialEM.|
|Gain Ref Policy||Opens dialog box to set policies governing multiple gain references.|
|View||Acquires image with first parameter set.|
|Focus||Acquires image with second parameter set.|
|Trial||Acquires image with third parameter set.|
|Record||Acquires image with fourth parameter set.|
|Preview||Acquires image with fifth parameter set.|
|Montage||Starts montage acquisition to file.|
|Prescan||Acquires binned-down montage without saving.|
|Set Minimum Counts||A submenu with settings to manipulate the filters used when correlating montage overlap zones.|
|Use Filter Set 2||Use the second set of filter parameters instead of the primary one.|
|Use Set 2 in Low Dose||Use the second set of filter parameters for montages taken in Low Dose mode.|
|Divide Filter Set 1 by 2||Divide each frequency of first set of filter parameters by 2.|
|Divide Filter Set 2 by 2||Divide each frequency of second set of filter parameters by 2.|
|Test Values for Set 1||Set values of first set of filter parameters for temporary use.|
|Test Values for Set 2||Set values of second set of filter parameters for temporary use.|
|Redo Corr when Read||Recompute correlations when reading a montage instead of using values from .mdoc file.|
|Set Minimum Counts||Set the minimum required mean counts for a montage frame.|
|Stop Acquire||Stops image acquisition.|
|Show Gain Ref||Shows the gain reference last used to normalize an image.|
|Show Dark Ref||Shows the dark reference last used.|
|Post-actions||Allows actions like tilt and image shift right after exposure.|
|Divide 16-bit by 2||Divides data from 16-bit camera by 2 in camera plugin.|
|Normalize Here||Toggles gain normalization within SerialEM on and off.|
|Interpolate Dark Refs||Interpolate between dark references at different exposures instead of taking one to match each exposure time.|
|Set Frame Align Lines||Set amount of detail (# of lines of output) in log from K2 frame alignment|
|Special Settings||A submenu with settings for test or specialized purposes.|
|Set Timing||Set timing parameters for dual-shuttering mode.|
|Set Quality||Set quality value for continuous mode images.|
|Set Scanning||Set parameters for scanning with a slit beam.|
|Set Drift Interval||Set interval for image shift changes when doing drift correction.|
|Set Corrections||Set basic image corrections performed in DigitalMicrograph.|
|Screen Down||Leaves screen down during image acquisition.|
|No Norm of DS Dose Frac||Turn off normalization of unnormalized K2 dose-fractionated images.|
|Simulation||Gets simulated images rather than acquiring from camera.|
|Debug Mode||Outputs debugging information to Log Window and Digital Micrograph Results window.|
Parameters command (Camera menu)
Use this command to open the Camera Setup dialog box and set parameters like exposure time, field of view and binning for the 5 different exposure modes.
Prepare Gain Ref command (Camera menu)
Use this command to open the Gain Reference dialog box and acquire a gain reference that will be used for gain normalization within SerialEM.
Gain Ref Policy command (Camera menu)
Use this command to open the Gain Reference Policy dialog box and set options controlling which gain reference is used for gain normalization within SerialEM when several are available.
View command (Camera menu)
This command acquires an image with the first camera parameter set. This would be a good parameter set to use for highly binned, continuous exposures, if continuous exposures worked reasonably well.
In Low Dose mode, images captured with these parameters are not shifted away from the center of the recording area. The View area is intended to be set up to give a low magnification, low exposure overview. The various procedures which would ordinarily lower the magnification to track positions, such as the eucentricity-finding routines, use View exposures in low dose mode. In other words, these routines rely on the parameter set giving adequate exposures for low-mag tracking in low dose mode.
Focus command (Camera menu)
This command acquires an image with the second camera parameter set. This parameter set is used by the Autofocus routine to acquire two or three images in quick succession, so it is typical to set it up for a small image that can be read out quickly, such as the central quarter of the field, with somewhat short exposure and drift settling times. It would be nice if one could also use this parameter set for visually assessing focus, but one usually wants a larger area for that purpose.
In Low Dose mode, images captured with these parameters can be, and typically are, shifted away from the center of the recording area.
Trial command (Camera menu)
This command acquires an image with the third camera parameter set. This parameter set is typically set up to acquire a full-frame image relatively quickly. It is used extensively by various routines that do tracking (e.g., eucentricity-finding routines). You should thus make sure that this parameter set gives an adequate exposure for tracking. Many of these routines actually compose a separate parameter set, changing the binning and exposure time if necessary so that binned images of no more than 512 pixels are acquired. However, when the tilt series controller acquires tracking images, it uses the Trial parameters without modification. It is recommended that you set these parameters to bin images binned down to no larger than 512 pixels, and that you acquire a full-frame image unless you have some specific reason not to.
In Low Dose mode, images captured with these parameters can be, and typically are, shifted away from the center of the recording area.
Record command (Camera menu)
This command acquires an image with the fourth camera parameter set. This parameter set is intended to be used for final acquisition of images to be saved, and the Tilt Series Controller will rely on it being set up for that purpose. In addition, montaging is done by acquiring images with these parameters.
In Low Dose mode, images taken with these parameters define the center of the recording area and can be considered unshifted.
Preview command (Camera menu)
This command acquires an image with the fifth camera parameter set. This is probably useful only in Low Dose mode, where it acquires an image from the Record area, using the same magnification and beam settings as for Record images. It can be set up to provide a very low exposure image of the Record area.
Montage command (Camera menu)
This command will start the acquisition of a montaged image, with the pieces saved to file. If there is not yet an open image file, you will first enter the Montage Setup dialog box, which allows you to specify the number of frames, magnification, and binning. From there, you enter the File Properties dialog box then the Save As dialog box.
Prescan command (Camera menu)
This command will acquire a binned-down montaged image, which allows a quicker assessment of the area that will be captured in an actual montage. The images are not saved to file. However, montaging needs to be set up first, which does entail defining an output file. If montaging has not been started yet, you will first enter the Montage Setup dialog box, then ensuing dialogs for opening a file.
Use Filter Set 2 command (Camera - Correlation Filter submenu)
The Correlation Filter submenu has a set of commands for controlling the filter used when correlating montage overlap zones. There are two sets of filter parameters available. Each set has default values, but the defaults can be changed by adding entries for MontageFilterR1R2S1S2 or MontFilterSet2R1R2S1S2 to the SerialEM properties file. The only difference between the defaults is in the low frequency filtering applied when 'Treat as very sloppy montage' is checked in the Montage Control panel. Filter Set 1 has a strong low-frequency filter, which has been found to be important when there are strong, high-contrast features like grid bars in the overlaps zone. However, this amount of low-frequency filtering can be detrimental when aligning low-dose montages without many features beside the regularly spaced holes. Filter Set 2 eliminates the extra low frequency filter and may work better for such specimens.
Use this command to switch to Filter Set 2 unconditionally. If you change filters and want to see the effect on alignment of an existing montage, you need to turn on the Redo Corr when Read option so that correlations are recomputed when the montage is read in.
Use Set 2 in Low Dose command (Camera - Correlation Filter submenu)
Use this command to switch to the second set of filter parameters for correlating montage overlap zones only in Low Dose mode. A low magnification grid map, taken outside of Low Dose mode, would use Filter Set 1, which may work better for montages full of grid bars.
Divide Set 1 by 2 command (Camera - Correlation Filter submenu)
Divide Set 2 by 2 command (Camera - Correlation Filter submenu)
Use these commands to divide all of the parameters of the first or second filter set by 2. The filter parameters are tuned to work with a moderate amount of noise in the images. When montaging in Low Dose mode, these parameters will work if either the dose or the binning is high enough (typically 4 for 4K camera). If taking images with an optimal binning for this situation is not feasible or not desired, then the filter parameters can be divided by to achieve an equivalent effect to doubling the binning. If you change this selection and want to see the effect on alignment of an existing montage, you need to turn on the Redo Corr when Read option so that correlations are recomputed when the montage is read in.
Test Values for Set 1 command (Camera - Correlation Filter submenu)
Test Values for Set 2 command (Camera - Correlation Filter submenu)
Use these commands to try different values of the filter parameters; your
settings here will not be retained between sessions. Parameters are all
expressed in cycles per pixel (Nyquist is 0.5). Entry boxes will come up
for each of the four parameters in turn:
Radius 1: The negative of the frequency at which the low-frequency filter starts to rise from 0 when the 'Treat as very sloppy montage' option is on; all frequencies below this radius will be set to 0. Only a negative value should be entered.
Radius 2: The frequency at which high-frequency filtering starts. This may need to be lower for very noisy data.
Sigma 1: The sigma for the low-frequency filter, which is an inverted gaussian that rises from 0 to 1.
Sigma 2: The sigma for the gaussian falloff of the high-frequency filter. This should generally be changed proportionally if Radius 2 is changed.
If you want to see the effect of new settings on an existing montage, turn on the following option.
Redo Corr when Read command (Camera - Correlation Filter submenu)
If you change any of the filter settings above and want to see how they work with an existing montage, you need to turn on this option so that the program will recompute the correlations with the new parameters when reading in the montage. Otherwise, it will use the shifts stored in the .mdoc file associated with the image file. In fact, with this option on, new shifts will be stored in the .mdoc file every time a montage is read in.
Set Minimum Counts command (Camera menu)
Use this command to set a minimum required number of mean counts for each montage frame. When a frame is taken that falls below this limit, the program stops and gives you three choices: to retake the image and test it again against the limit, to save the image and go on, or to stop the montage. However, if the property ShootFilmIfMontageDark is set to 1, then the program will first take a film picture, which may unstick the lower shutter on a JEOL. Set the minimum counts back to 0 to disable this test.
Stop Acquire command (Camera menu)
This command will stop image acquisition if it is still in some intermediate state, such as raising the screen or acquiring a dark reference. However, once the request for an image has been issued, stopping will have no effect.
Show Gain Ref command (Camera menu)
This command will place the gain reference that was last used to normalize an image into buffer A and roll the contents of A into B, etc., to the extent that buffer rolling is selected. The gain reference consists of real numbers around 1.0 that are multiplied by the image to normalize it.
Show Dark Ref command (Camera menu)
This command will place the last-used dark reference into buffer A and roll the contents of A into B, etc., to the extent that buffer rolling is selected.
Post-actions command (Camera menu)
This command toggles a mode in which various time-consuming actions are initiated immediately after camera exposure ends, at the beginning of the image readout. These actions can be magnification changes, image shift, and stage tilting. Stage tilting is the most noticeable action, because it means that when you stop the Tilt Series Controller, it will often have already gone on to the next tilt. If this behavior is problematic, you can disable it with this command.
Divide 16-bit by 2 command (Camera menu)
This command toggles a mode in which all images from a 16-bit camera are divided by 2 in the camera plugin, prior to being received by SerialEM. In this mode, negative numbers in a dark-subtracted or gain-normalized image will be preserved instead of truncated at zero. This could be important for low-exposure images if the dark reference drifts downward. Another advantage of this mode is that the numbers shown in SerialEM will match those saved to a file. When this mode is selected, the options for treatment of 16-bit data in the File Properties dialog box will be disabled because they are no longer relevant. (The image has already been divided by 2 and will not be recognized as an unsigned 16-bit image.)
Normalize Here command (Camera menu)
This command toggles the processing (dark subtraction and gain normalization) of images within SerialEM. This setting is initially determined by the global 'GainNormalizeInSerialEM' property or by the 'NormalizeInSerialEM' property for individual cameras. When it is off, processed images are acquired from DigitalMicrograph (or TIA for an Eagle camera). When it is on, raw images and dark references are acquired from DigitalMicrograph, and the processing is done in SerialEM. In the latter case, the gain reference could come from DigitalMicrograph or from one prepared in SerialEM, depending on which exists.
Interpolate Dark Refs command (Camera menu)
This command toggles whether the program will compute a dark reference for a particular exposure time by interpolating between dark references for two other exposure times, instead of taking a dark reference to match each exposure. When this feature is enabled, interpolation is used if the desired exposure time is between the times for two existing references separated by no more than 0.2 second. If such references do not exist, a new reference is taken either at the desired exposure time or at an interval of 0.2 seconds from a nearby reference; in the latter case interpolation can then be used. The interpolated dark reference will be at least as accurate as a reference taken to match a particular exposure, because interpolation averages over two estimates of the reference. Thus, it is not necessary to take averaged dark references to get good interpolated references, although averaging will improve the estimate further.
When exposure time is being varied automatically during a tilt series, the program activates interpolation if possible. However, this feature is available only when gain normalization is being done in SerialEM.
Set Frame Align Lines command (Camera menu)
This command allows you to control how much detail is printed to the log window after K2 frame alignment done in the SerialEMCCD plugin to DM. The default is two lines of output: one with the raw and smoothed movement distance implied by the frame shifts; one with selected residual values. If enough frames are included when fitting the shifts to allow robust fitting, the residuals given are the average of the weighted mean residuals of the fits (an overall figure of merit), the maximum over the fits of the maximum weighted residuals in each fit (reflecting the worse frame alignment result), and the maximum of maximum unweighted residuals(reflecting the occurrence of bad alignments that could be discarded as outliers). With no robust fitting, it simply shows the overall mean residual and maximum of maximum residuals.
When you select to show one line, it will show the overall mean residual and maximum of maximum residuals, based on weighted values if there is robust fitting, plus the raw distance moved.
When you select to show three lines, it will compute the Fourier ring correlation (FRC) between sums of even and odd frames and report the frequencies values at which it falls below various levels, plus the FRC at half-Nyquist.
Set Timing command (Camera menu)
This command allows you to enter values for the startup delay, minimum drift settling, and extra beam time needed in the dual shuttering mode of operation. The option is enabled by being in Administrator mode.
Set Quality command (Camera menu)
This command allows you to enter a value for the quality level to set when acquiring images in continuous mode from the current Gatan camera. Enter a number between 1 and 7 (most camaeras will not have that many quality levels), or enter 0 to have no quality value set.
Set Scanning command (Camera menu)
This command was used to enter parameters to control beam movement and focus changes while scanning with a slit beam.
Set Corrections command (Camera menu)
This command allows you to enter a value for the basic corrections to be done when DigitalMicrograph acquires an image. This value may initially be set with the 'BasicCorrections' property, otherwise it is -1, meaning that default corrections will be done. The value should be the sum of 1 for defect correction, 16 for bias correction, and 32 for linearization. 0 will turn off all default corrections. This command is enabled by being in Administrator mode.
Set Drift Interval command (Camera menu)
It is possible for drift to be compensated during an exposure by using a script command to set the drift rate for the next exposure. This command is used to adjust the interval between changes in image shift during the exposure. The default is 100 ms, which may be too short on a JEOL.
Screen Down command (Camera menu)
This command is used to toggle a mode in which the screen is not raised before a camera exposure. This could be useful for looking at the beam during an exposure.
No Norm of DS Dose Frac command (Camera menu)
This command can be used to turn off the normalization of the summed image that is returned from a K2 dose fractionation shot taken without gain normalization.
Simulation command (Camera menu)
This command is used to toggle simulation mode, in which fake images are produced within SerialEM rather than acquired from DigitalMicrograph. It should not be needed for simulation if DigitalMicrograph is started with Ctrl and Shift held down to activate the Faux camera.
Debug Mode command (Camera menu)
This command is used to toggle debug mode, in which the SerialEMCCD plugin will chronicle its transactions in the DigitalMicrograph Results window. The camera controller in SerialEM will also output some timing information in the Log Window. After turning on this mode, you can debug problems with taking a picture with the following procedure:
Be sure that the Results window is open in DigitalMicrograph.
Try to take an image of the kind that makes an error occur.
Go to the DigitalMicrograph Results window. At the end will
be the last script executed. Open a new script window with Ctrl-K. Copy from
the Results window, starting at the line
Object manager =...
after the last
DMcamera executing script :
up to the line before
Paste this into the script window. Run the script with Ctrl-Enter. It should give an error message with more details about what is bothering it. If the solution is not obvious, copy down the error message, save the Results window, and send them to David Mastronarde.