problem with Gatan camera and possible issues for Leginon
Added by Anonymous about 19 years ago
One of the microscopists noticed that there's a problem with our Gatan camera. This may be due to the FEI Tecnai microscope working together with the Gatan camera less than perfectly. The microscopist noticed that in low dose mode there appears to be a double exposure occurring. When he disconnected the cable for the screen lift, he saw that there's a double flicker occurring when attempting to take a picture with the Gatan camera - implying that the specimen is exposed to electrons twice for a single micrograph. So possibly the screen blanking or something is not working correctly. This could be a problem for obtaining two images (e.g. a different defocuses or tilt) from a single area; because it will already have been exposed to too much radiation damage for the second image. I'm wondering if this is going to be a problem for Leginon. We generally run Leginon so it takes an image at a near focus exposure preset and another image at a far focus preset. I was wondering if Leginon obtains these images in a sufficiently different manner that it can avoid the problems (due to the microscope and camera apparently working incorrectly) affecting manually obtaining pairs of images in low dose mode,
William
Replies (7)
Re: problem with Gatan camera and possible issues for Legino - Added by Anonymous about 19 years ago
"WVNicholson" wrote: One of the microscopists noticed that there's a problem with our Gatan camera. This may be due to the FEI Tecnai microscope working together with the Gatan camera less than perfectly. The microscopist noticed that in low dose mode there appears to be a double exposure occurring. When he disconnected the cable for the screen lift, he saw that there's a double flicker occurring when attempting to take a picture with the Gatan camera - implying that the specimen is exposed to electrons twice for a single micrograph. So possibly the screen blanking or something is not working correctly.
Sorry, I think I meant to say beam blanking rather than screen blanking.
"WVNicholson" wrote: This could be a problem for obtaining two images (e.g. a different defocuses or tilt) from a single area; because it will already have been exposed to too much radiation damage for the second image. I'm wondering if this is going to be a problem for Leginon. We generally run Leginon so it takes an image at a near focus exposure preset and another image at a far focus preset. I was wondering if Leginon obtains these images in a sufficiently different manner that it can avoid the problems (due to the microscope and camera apparently working incorrectly) affecting manually obtaining pairs of images in low dose mode,
William
Gatan camera - Added by Anchi Cheng about 19 years ago
Hi, William,
Christrain might be the best to answer your question in full. However, here is my input.
Leginon controls Gatan camera through Digital Micrograph. The trouble shooting session in the Leginon Manual describes the correct configuration
within Digital micrograph. There are several ways for Gatan camera to take exposure. One uses only the primary shutter, one requires two signals that goes to the two shutters. Leginon uses the former. The flicker your user observes suggest that they may be using the latter. To check if the proper shutter recieves the signal, observe the leds on the Gatan camera control box front panel. Of course you will first need to know which shutter cable is connected to 1(primary) and which to 2. Compare that with what you observes on the main screen of the TEM (Leginon does not requires screen up when taking CCD images. Therefore, you can leave the screen down to see the beam coming and going), then you will know whether Leginon behaves correctly.
If the cables and configurations are matched correctly, you should not have double exposure problem in Leginon.
Re: Gatan camera - Added by Anonymous about 19 years ago
"anchi" wrote: Hi, William,
Christrain might be the best to answer your question in full. However, here is my input.
Leginon controls Gatan camera through Digital Micrograph. The trouble shooting session in the Leginon Manual describes the correct configuration
within Digital micrograph. There are several ways for Gatan camera to take exposure. One uses only the primary shutter, one requires two signals that goes to the two shutters. Leginon uses the former. The flicker your user observes suggest that they may be using the latter. To check if the proper shutter recieves the signal, observe the leds on the Gatan camera control box front panel. Of course you will first need to know which shutter cable is connected to 1(primary) and which to 2. Compare that with what you observes on the main screen of the TEM (Leginon does not requires screen up when taking CCD images. Therefore, you can leave the screen down to see the beam coming and going), then you will know whether Leginon behaves correctly.If the cables and configurations are matched correctly, you should not have double exposure problem in Leginon.
I found the relevant section of the Leginon manual which says that the configuration in Digital Micrograph should have in the camera configuration the Primary Shutter set to Pre-specimen, the Alternate Shutter set to Post-specimen and the Default Primary Shutter set to the Normally Closed position. However, I talked to Allen Boswell from FEI and he seemed to think we should have the Alternative Shutter set to Pre-specimen, the Primary Shutter set to Post-specimen and the Alternative Shutter set to the Normally Closed position presumably because our microscope and camera are set up differently. Does that sound correct? Also, would there be any advantage in swapping the cables round on the camera control box to allow having the Digital Micrograph settings exactly as they appear in the manual?
William
Shutters and Gatan Camera - Added by Anchi Cheng about 19 years ago
Hi, William,
First I should make a correction of my earlier statement. Leginon does not allow shuttering with screen down unless, as you mentioned, that the override cable is unpluged from the camera control box.
Leginon uses a single shuttering mechanism. Therefore if you put the pre-specimen shutter cable on the alternative shutter line (position 2) and set in the software to control the shuttering through alternative shutter normally closed, you should get the same result as doing all these on the primary shutter. The main question is where the pre-specimen shutter such as the beam blank shutter is connected to. If you have other users in the facility who need to use the double shuttering such as tomography pre-exposure, it may be better to be consistent in cable connection. We recommend putting pre-specimen shutter on primary only because that is how we configure it and it has worked.
Gatan 4k camera bias level and quadrant reference - Added by Anchi Cheng about 19 years ago
Here is quote from the reply email by Brent Bailey of Gatan (bbailey@gatan.com ) about bias level and quadrant reference. It explains how these two corrections are made and confirms my earlier posting about the raw (unprocessed) image in Leginon were bias-stabilized and quadrant-corrected.
" Each image is read out with an extended pixel region, that is used to measure the instantaneous DC level of the video signal in the absence of light. In your case, each quadrant is 2048 pixels wide. Each time a line is read out from a quadrant, we clock out additional pixels after we get to 2048. Since the serial resister is empty after the 2048th pixel has been clocked out, the extended pixels give the video level at that point in time in the absence of signal. This is important because the level will drift over time and needs to be constantly monitored.
The "Set Bias" function tells the camera to monitor the extended pixel region and apply sufficient external bias to each video output through a set of digital to analog converters (DACs) so that the extended pixel region is at 1000 counts nominally. This is done to maximize the dynamic range of the camera while giving enough of an offset so that the A/D converters never get down to 0 counts which would result in truncation of data. Normally Set Bias only needs to be run during installation. The values are stored in the "Preferences" file and loaded into the controller each time Digital Micrograph is run. If there is a problem with the bias levels you may see a dark quadrant(s). In older versions of Digital Micrograph, there were cases reported in which the correct values were not loaded properly during startup. In this case either quitting DM and starting again or running Set Bias usually corrects the problem. This issue is corrected in current versions of DM. Once the bias levels of the 4 extended pixel regions are set, DM automatically subtracts an average level from each extended pixel region from its associated quadrant. The software then adds 1000 counts to the difference so that there are no negative values. The system is now bias stabilized.
Quadrant correction is a proprietary algorithm that applies appropriate scale factors to images to suppress the visibility of quadrants. In a uncorrected system, quadrant visibility occurs because of small deviations from linearity in the four CCD outputs. Once "quadrant correction" has been run "unprocessed" DM images are actually bias-stabilized and quadrant-corrected. The quadrant correction scale factors are stored in DM until the algorithm is run again at which point they are overwritten. Since you are using "unprocessed" images to get your dark and gain reference images, you are actually using a method similar to the built-in DM dark subtracted or gain normalized acquisition modes.
In order for quadrant correction to work properly it is important that uniform illumination be used both during the quadrant correction algorithm and during normal operation. Heavy shading in an image due to poor beam alignment may result in quadrant visibility."
"Location" in DM CCD camera configuration? - Added by Anonymous over 16 years ago
By the way, what option do you recommend for "Location" in DM CCD camera configuration? The Leginon manual does not appear to have a recommendation for "Location". The closest I can find to a description of "Location" in the Gatan CCD camera manual is that it relates to the user viewing images in a postshutter location (GIF, TV screen, etc) and can affect how shutter override behaves (so if that explanation is correct then the option chosen for "Location" is important). Unfortunately, when I asked our Gatan service engineer what "Location" is he said that is unimportant. I can't decide which option to pick out of "None", "High Resolution Electron Microscopy" or "Unspecified". There isn't a description of these options for "Location" in Gatan's manual. I'm currently using "None" as that seemed to work in the past but it may not actually be the best setting?
William
- Added by Anchi Cheng over 16 years ago
William,
I have no idea what Location Option is for. I assume that in some case there are more than one camera and you can pick which one to use for focus different from the one to use for record the final image.
Since we have only one camera, there is no such an issue. If you have more than one camera and use different ones for focus and recording when you are on DM, and you are unhappy with how Leginon does manual focusing using the recording camera, I suppose that you can add your other camera to your instrument list and use that for manual focusing by choosing that camera in fc preset settings. You are on your own if you want explore this as we can't try it here.
For the record, our DM comera configuration is set for "High Resolution", although I think it does not matter because there is only one camera.
Anchi