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Serials6pm; Already have a WordPress.com account? Serials6pm; Customize; Follow Following; Sign up; Log in; Report this content; Manage subscriptions. Automated EM Data Acquisition with SerialEM. Tilt Series Acquisition with SerialEM Single Particle Acquisition with SerialEM Program Features Contribute to the Continuation of SerialEM.

Automated EM Data Acquisition with SerialEM


Tilt Series Acquisition with SerialEM
Single Particle Acquisition with SerialEM
Program Features
Contribute to the Continuation of SerialEM
Highlights of SerialEM 3.8
Availability of SerialEM for Various Hardware Configurations
Setting Up SerialEM
How to Report a Problem with SerialEM
SerialEM Discussion Group
Other Helpful Materials
Source Code for SerialEM and SerialEMCCD Plugin
Download and Installation Page

Tilt Series Acquisition with SerialEM

SerialEM is a program that can acquire a variety of data fromelectron microscopes: tilt series for electron tomography, large image areasfor 3-D reconstruction from serial sections, and images forreconstruction of macromolecules by single-particle methods.

The main focusof the program is automated tilt series acquisition. For this, it uses an approach based on prediction of specimen positionduring the tilt series from the position at previous tilts. It does not counton the microscope or the specimen being particularly well-behaved, so unless aprediction appears reliable, it falls back to measuring and adjusting defocusand/or specimen position when necessary. With this method, it achieves bothrobustness and speed. For example, a2Kx2K, 1 degree tilt series can be acquired in about 20-25 minutes with a four-port readout CCD camera.

The method and the program are described in this paper:
Mastronarde, D.N. 2005. Automated electron microscope tomography using robustprediction of specimen movements. J. Struct. Biol. 152:36-51.
The prediction method was first described in thisabstract from the 2003 MSA meeting,
Mastronarde, D.N. 2003.SerialEM: A program for automated tilt series acquisition on Tecnaimicroscopes using prediction of specimen position. Microscopy andMicroanalysis Vol. 9, Suppl. 2, 1182CD.

Single Particle Acquisition with SerialEM

SerialEM is also widely used to acquire data for single particlereconstruction. After positioning on a hole, it can acquire multiple image ina hole and/or in multiple holes using image shift, with compensation for thebeam tilt induced by the image shift. There is a routine that finds holepositions automatically and one that can combine them into an optimal subset ofpositions for acquiring from multiple holes. The basic framework for thisautomated acquisition is through the Navigator module, but there is currentlyno user interface for the various additional options needed for efficient,high-quality data acquisition. Such options are provided by SerialEM'sscripting capability, and there are various sophisticated collections ofscripts available in the EM community.

Program Features

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In addition to automating data acquisition, the program provides anintegrated user interface for image acquisition, display, and storage. Itlooks like this:


Click image for full-sized view.

Other notable features include:

  • The program provides a flexible interface for user intervention.When acquiring a tilt series, the user can stop if necessary, adjust position or focus, and even conveniently back up to a previous tilt angle and replace the stored data from that point onward. Other kinds of acquisition also provide an ability to stop and restart.

  • There is a low dose mode, in which focusing and tracking are done in aseparate area to minimize exposure of the area being recorded. The programallows one to set up the magnification and illumination independently for upto 4 different areas, where these settings are invoked simply by taking apicture of the particular area.

  • SerialEM can automatically acquire a montage of overlapping frames, and itcan acquire a tilt series of such montaged images.The images are written to a single file that can be viewed immediately in3dmod. A montaged tilt seriescan be processed with the eTomo interface in IMOD.

  • SerialEM can also acquire energy filtered images through a Gatan ImagingFilter (GIF) or a JEOL omega filter. It implements several features to makethe GIF more convenientto use, such as automatic switching between regular magnifications with theviewing screen down and the EFTEM lens series needed to use the GIF with thescreen up. Both low dose mode and montaging can beused with the filtering.

  • The program supports imaging and tilt series acquisition in STEM mode.

  • There is a Navigator window that allows one to store and return tospecimen positions, make low-magnification maps, mark positions on maps, return to marked positions reliably, and automatically acquire images, tilt series, or montages at a series of positions.

  • The program has a scripting capability that includes variables, 1D and 2D arrays, simple arithmetic statements, looping, and conditional statements. It is most commonly used to acquiring images automatically for single-particle reconstruction; some scripts are available for this purpose.The script capability also provides a relatively easy way to add commands requested by users for specialized or experimental use.

  • The operation and configuration of SerialEM is completely documented in aWindows compiled HTML help file opened from the program. You can access the help online in HTMLformat for either the current stable release version or the development version of SerialEM. In addition, you can download and view the compiled help file for the development version after unzipping on any Windows machine.The on-line help and compiled file for the development version are continually updated and will often show features that are not inthe current release version.

Contribute to the Continuation of SerialEM

SerialEM was originally developed with grant funding from NIH.Since 2014, it has been maintained and developed with funding that comes fromusers, which has allowed us to focus on coding and documenting instead of grant-writing. Since 2018, funds have also been usedto pay Guenter Resch to provide user support.

You can helpmake SerialEM a community-supported resource by having your laboratory ormicroscope facility contribute to the further development and continuedavailability of the program. The mechanism for doing this is to purchasepriority annual support, so that the expense represents a tangible item that can bepaid from grant or institutional funds. In September 2020, we transitioned tohaving these payments made to Guenter Resch's company, Nexperion. In additionto directly funding Guenter's assistance to users, themajority of the funds are used to compensate David Mastronarde for his work onSerialEM.We offer threeoptions, allowing you to pick the size of your payment based on yourfinancial resources and the importance of SerialEM to your work. Theoptions are:

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  1. Basic support: Priority resolution of issues arising from SerialEM bugs andconsultation of issues arising from configuration and calibration problems($550 non-profit, $1100 commercial).
  2. Software support: In addition to basic support, advice and consultation on howto optimally use the workflows available via SerialEM’s user interface ($1100non-profit, $2200 commercial).
  3. Application support: In addition to software support, advice and consultationon how to get the most out of SerialEM for complex applications, e.g. viascripting ($2200 non-profit, $4400 commercial)

What happens if you do not pay? Although the first optionrepresents the kind of support that we provideroutinely, payment for this support is voluntary. We doanswer questions and deal with problems from non-paying users, but may takelonger to do so.

Why is it important to contribute? Usercontributions are important both financially and because they are not tied tospecific developments and represent the general user interest.

What does the change in 2020 mean for the future of SerialEM? This is not a movetoward commercialization. We intend to keep the program freely available. Itis already open source.

How do I get help with SerialEM under the new arrangement? For everyone, requests for support and for initial configuration (framework) files should still besent to [email protected] (See How toReport a Problem with SerialEM).

Click Here to go to the registration page for supportagreements on Nexperion's web site. (This initial registration is requiredonly once.)

Highlights of SerialEM 3.8

The most notable features of the latest release, SerialEM 3.8, are:

  • Dose-symmetric tilt series run from the tilt series controller, with options for flexible grouping of tilt images.
  • A procedure for finding a regular array of circular holes in a support film.
  • A procedure for converting a regular array of Navigator points into an optimal subset of points for acquisition from multiple holes using image shift.
  • Ability to take frames for saving and/or aligning from Tietz, OneView, and Rio cameras.
  • Options to return camera images in floating point or with extra divisions by 2 to avoid saturation from Tietz, OneView, and Rio cameras.
  • Fast continuous mode acquisition from Tietz cameras.
  • New integrated tasks and associated dialogs for conditioning a phase plate and waiting for drift to settle, and a new dialog to set parameters for CTF fitting.
  • Ability to save a snapshot of the image display window and items drawn on it, either the same size as on the screen or with selected zoom.
  • Increased scripting capabilities, with 109 new script commands.

There are many other improvements; see the list of notable features to learn about them.

Availability of SerialEM for Various Hardware Configurations

SerialEM works on modern Thermo Fisher (FEI) microscopes (Tecnai, Titan,Spirit, Talos, Glacios),JEOL microscopes with the TEMCON or TemCenter interface (e.g., 1400, 2100,2200, 2500, 2800, 3100, 3200, ARM200, F200, CryoARM), and the Hitachi HT7700 and HT7800.The program is built and tested primarily with a Tecnai simulator on Windows 10,but itstill works on Tecnais under Windows 2000.

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SerialEM works with CCD or CMOS cameras from Gatan, TVIPS (Tietz Video andImage Processing Systems), Thermo Fisher, AMT, DirectElectron, and EMSIS (formerly OSIS, Olympus Soft Imaging Solutions); it also supports the JEOL Rubyand other cameras. Direct detection cameras fromGatan, Thermo Fisher, and DirectElectron are supported. The EIGER2-based EMdetectors from DECTRIS are also supported through a plugin currently availablefrom DECTRIS.All EMSIS cameras now workthrough a plugin available from EMSIS for either 32- or 64-bit versions. STEMacquisition is supported from Gatan's DigiScan, Thermo Fisher's STEM imaging, and theJEOL STEM on the 1400 series.

SerialEM cannot control frame-saving from Falcon 3 cameras without a new 'advanced' scripting interface. This interface is not being provided for Tecnai/Polara microscopes. Contact Thermo Fisher for details, including whether a license will be needed and how to obtain it.

Status of K3 Camera Support: SerialEM's remaining issues with the K3 camera are listed inRemaining Problems with the K3.

If necessary or advantageous, SerialEM can be run on a separate computer andcontrol the microscopeover the network. Such a configuration is standard on JEOL microscopes, is quitecommon when using K2 and DE cameras on Thermo Fisher microscopes, and isalso possible for Hitachi microscopes. Version 3.5 of the program canaccess Gatan, Direct Electron, Thermo Fisher, and JEOL Ruby cameras overthe network from another computer, and version 3.6 can access Tietz camerasremotely as well, but for other kinds of cameras, SerialEMmust run on the computer that hosts the camera.

For running on a Thermo Fisher microscope, it is necessary to have the Thermo Fisher scriptingadapter installed. If you do not have the adapter, you need to purchase itfrom Thermo Fisher. To check whether it is installed, do the following:

  1. Locate and run the 'Delphi Scripting Exampler'. For Tecnais through version 3 and possibly on older Titans, this is available through the Start menu under Programs - Tecnai. For Tecnai version 4 and newer Titan software, it is under Programs - FEI Company - TEM Software.
  2. Select the 'Projection' tab after the program starts.
  3. Press the arrow next to 'Magnification'. If a magnification appears, you have the adapter; if not, there will be an error message in the status bar at the bottom of the window.

Although SerialEM itself is open source, the microscope plugins and somecamera plugins in this software are provided free for non-commercial purposes.By downloading the distributed software, you agree that you will use it forresearch rather than commercial purposes, and that you will not distribute itoutside of your own institution.

The program has been installed on over 300 Thermo Fisher and 200 JEOLmicroscopes.

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Setting Up SerialEM

Program and configuration files are available on the Download and Installation page. Follow theinstructions in there to get started. As a first step, fill out the questionnaire in Word doc form and send it to [email protected];we will prepare an initial configurationfile. This is strongly recommended for TFS/FEI scopes and absolutelyessential for JEOL and Hitachi scopes.Installations on new JEOL microscopes are usually done with the assistance of Jaap Brink from JEOL(jbrink at jeol dot com), but sending in the questionnaire is still the starting point.

Guenter Resch from Nexperion offersSerialEM installation and training for both Thermo Fisher and JEOL microscopes. Thisservice is available worldwide, but visa arrangements for the the United States may need extra attention.Chen Xu at the University of Massachusetts Medical School and Gabriel Frank atBen Gurion University in Israel may alsobe willing to help with installation; email
for contact information.

How to Report a Problem with SerialEM

Send all problem reports and requests to [email protected] This is a private email address, not amailing list.Guenter Resch is now the primary contact for getting assistance withSerialEM. He will collect needed information about a problem, and handle mostproblems with use of the program. Problems in the program itself will behandled by David Mastronarde.

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The following information and files can help to address your question fasterand more effectively. Here are the essentials:

  • The exact wording of the error message
  • A saved log if it might contain relevant information, especially the logafter a crash. When the program crashes, it saves to the log file ifone was defined, otherwise to a file SerialEMcrash.log (or SerialEMcrash1.log,etc), typically in the directory where the initial settings file was located.
  • Screenshots of error messages, SerialEM, and/or the microscope userinterface. Please send a real screenshot of the whole SerialEM window andassociated components. We find that cell phone pictures are often unreadable,and that some useful information is often just outside the range of a partialscreenshot.
  • A precise description of steps that lead to the issue you are reporting
  • How often the problem occurs (every time, sometimes, about once a day)
  • The type of microscope and its location (city, institution, and laboratory).
  • The type of camera being used when the problem occurred.
  • The exact version of SerialEM, including whether it is 32- or 64-bit, and fora beta version, the build date. This information appears at the start of thelog when the program starts, and as of SerialEM 3.7 it is in the crashmessage.

These additional points can also be helpful:

  • SerialEM properties and calibrations (SerialEMproperties.txt andSerialEMcalibrations.txt, typically found in C:ProgramDataSerialEM orC:Program FilesSerialEM). Also describe which directory the files came from.
  • If the properties file is not available when you report, describe the layoutof your system (what hardware is used, what computers are present, where isSerialEM installed)
  • The settings file that you were running from. (If not a specially named file,right-click the shortcut that you run SerialEM with, select properties andfind SerialEMsettings.txt in the 'Start in' folder shown there.)
  • The version of the operating system, including whether it is 32- or 64-bit,and of the microscope user interface being used
  • Whether you tried any of the relevant troubleshooting procedures described inthe SerialEM help, and if so which ones.
  • If the program hangs, for how long did you let it sit before killing it?

Please follow these additional tips for effective communication:

  • Do not reply to a previous message from the SerialEM list or one of us;start a fresh message with its own title line.
  • Report one issue per email unless there is some apparent relationship betweenthe issues.
  • Follow this terminology for the behavior:
    • Crash: the program closes, generally after an uninformative error message from SerialEMand/or Windows.
    • Hang: the program becomes unresponsive and has to be closed with Task Manager.
    • Error: a procedure stops with an error message from SerialEM
    • Bad state: the program interface is responsive but many operations are grayedout and pushing STOP does not restore it.

SerialEM Discussion Group

The SerialEM discussion group provides a means for users and peoplemanaging the program to ask questions and share their experiences, as well asa way to receive notification of new releases and other news.Digests and posting via email (i.e. list server functionality)are provided. Additional capabilities, including browsable / searchablearchives are available through a web interface.Access to the web interface requires a Google login profile,which must be associated with the email address at which you wish tobe contacted. (Non Gmail addresses can be used with a Googlelogin, with or without a Gmail address. See the Email section of yourGoogle Account settings).The postings are not moderated, but joining requires approval.

To subscribe or visit the group via the web interface please visit SerialEM.If you do not have and are not willing to create a Google login and are satisfied with email only access, you can also request membership by sending an email to
asking to be added to the SerialEM list, and you will be added manually. Be sure that this email is sent from the same address as youwould use to post to the list.

To cancel your subscription, use the web interface orsend an email to [email protected]

When replying to a previous posting via email, you will have the optionto reply to the sender or to the list.

Other Helpful Materials

A standalone copy of 3dmod and associated files that can be run on Windows without installing IMOD or Cygwin is availablehere. The package unpacksto C:Program Files3dmod by default and a shortcut can be set up to run 3dmod fromthere. The package also includes midas, which can be useful to have on themicroscope for checking rotations between magnifications.

A repository for contributed scripts was developed by Guenter Reschfrom Nexperion and is hosted onthe Nexperion site. If you have scripts that may be useful to others, pleaseconsider adding them to the repository.

Daniel Caujolle-Bert at the University of Basel has written a program formanaging user and settings and scripts in an environment where multiple users sharethe same login; it is available here.

Our YouTube channel has a seriesof videos of the lectures on SerialEM presented in a webinar in September2012, plus some tutorial videos prepared by Cindi Schwartz. The videos arealso available right here; if you sort on the 'Last modified' column they will be in a goodorder for viewing.

Here are some training documents written by Cindi Schwartz, to provideconcrete guidance for doing room temperature and cryo tomography.

Here are Powerpoint presentations from a workshop on SerialEM given inDecember 2006. The slides have a lot of words, but there are many fewerwords than in the help manual, so they may be helpful by highlighting the mostimportant points.

And again, here are the:

Source Code for SerialEM, SerialEMCCD, and Python Plugins

Both SerialEM and the SerialEMCCD plugin to DigitalMicrograph are open source;SerialEM under an MIT license, and SerialEMCCD under the GPL version 2. You canbrowse the SerialEM source here and SerialEMCCD source here, or you can download eitherrepository with Mercurial. Seethe IMOD source instructions for setting up Mercurial and updating a repository. ThisSerialEM repository has been pruned of all changes before Feb. 17, 2017 so asnot to disclose calls to proprietary microscope and camera interfaces.However, it is updated nightly from the main development repository, withchanges committed with the original comments.

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To clone the open-source SerialEM repository, run

To clone the SerialEMCCD source, run

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If you are going to try to compile either of these projects, you will alsoneed the collection of libraries, which is now in a separate SerialEMLibs repository. You can download this with:

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The module for Python support is also available here. You can browsethe project files here or you can clone thisrepository with: