Table of Contents
Frequently Answered Questions
If you have questions that are not covered below, please consult the
on-line documentation.
Contact
support staff
for additional
questions. During your beamtime, use the beamline cell
phones.
In Blu-Ice, click on the units next to the energy input box to
toggle between wavelength (Å) or energy (eV or keV).
The backstop should be placed at a position where it allows collection
of reflections in the 30-40 Å resolution range. At most
wavelengths, it will be possible to collect even lower resolution,
however, this will be at the expense of additional air
scatter that may obscure weak reflections and reduce the
diffraction signal
over noise.
The Blu-Ice resolution predictor shows the
low resolution limit at the given beamstop and energy values.
The white box in the sample camera video displayed on
Blu-Ice represents the approximate FWHM beam size at the sample position at
all the zoom levels
(unlike the box displayed in the monitors at the beamline, which does
not change size with the camera zoom. With the on-axis camera,
the box shows what parts of the sample to beam hits. With the
orthogonal sample camera, the vertical dimension of the box shows the
size of the beam at 90 degrees.
No, this is normal operation. The system uses the medium zoom level to do the alignment,
in case large loops are used.
Also, during the crystal screening mode, maximum zoom is used for
recording JPEG images of the crystal.
You can view the image header in ADXV.
The default beam position,
should usually be at the nominal centers of the detectors (within 1 pixel) listed below (in mm):
- Pilatus 6M: 211.8, 217.3
- Eiger 16M: 155.6 163.9
When a detector offset is selected in the Blu-Ice hutch tab, the data collection software writes the
true beam center coordinates Cx,Cy to the image header.
Note: For CCD detectors and the Pilatus 6M, The convention used at SSRL may not be the same as on other
sites. If the detector is offset and you cannot process the data, try
flipping the Cy coordinate as described below.
Pilatus and Eiger detectors
To get the best data, we recommend 0.1 or
0.2 degrees oscillations and short exposures per image. However,
a larger oscillation (0.5 -1 degree) should be used for crystal
screening or test shots. If in doubt, use the default parameters in
Blu-Ice .
Important: Thin sliced images with the correct exposure time look very weak
- this is to be expected, as all the reflections are partials, and
you should refrain from increasing the exposure time until the
pattern looks strong, since this may result in rapid radiation
damage. Even if you do not exceed the maximum allowed dose with an
increased exposure time, consider increasing the data multiplicity
instead.
Small oscillation data can be processed at SSRL in a very straightforward
manner either with XDS (see also the SSRL script
autoxds)
HKL3000 and the most recent version of MOSFLM can be
also used, although we have found that XDS tend to give the best
results for oscillations much smaller than the crystal mosaicity.
Because both the Pilatus and Eiger detectors have a very short readout time compared with
the typical exposure time, it is not necessary to stop the phi
rotation or close the shutter while the detector reads out the
image; this results in faster data collection and, for very short
exposures, may reduce systematic errors. The default setting
for data collection mode is to collect the entire data set without
closing the shutter or stopping phi.
For MAD and SAD data collections the shutter will close and phi
will stop at the end of each wedge to change the energy or the crystal
orientation. For native data sets, entering a wedge value different
than the default of 180 will also cause the shutter to close,
dividing the data collection run into several shutterless
intervals. This can be advisable to be able to pause the data
collection at some point - for instance, in order to check the preliminary results of
data processing (see following section)
Choosing a wedge equal to the oscillation range effectively turns
off shutterless data collection. This is not recommended, with the
possible exception of MAD or SAD experiments on crystals that suffer
significant radiation damage after a single shot.
The ''Pause'' button in the Blu-Ice data
collection tab is designed to wait until the shutter closes (so that
the last image is not bad). This means that after pressing this
button, data collection will continue until the end of the
current wedge.
Similarly, if there is a SPEAR3 current dump in the middle of the data
collection, the collection will not pause, but proceed to the end of
the wedge. Multiple images will be blank. For this reason, it is
important to always monitor data collection with the Pilatus or Eiger
detector during shutterless data collection- or use
wedges small enough that can be recollected without increasing
dramatically the total dose absorbed by the crystal.
The Abort button can be used to stop the data
collection, although a few images will be collected before the
command takes effect. If you wish to resume the data collection after
the pause, remember to recollect the last image written to disk.
The Pilatus detector is programmed to change the gain at an energy of
9000 eV. Setting the new gain for all the pixels can take about two
minutes and this will happen every time you start a data collection
run at an energy that crosses that threshold in either direction:
eg. if you collect at 12000 eV on one run and 8000 eV in the
next run or vice-versa. If you want to do a MAD experiment on an
absorption edge below 9000eV and the remote energy is above 9000eV
consider manually selecting a different remote energy. Ask the
support person for advice.
While the Eiger detector takes a shorter time to reset the gain
(about 20 seconds), it does this over a shorter energy change, and
it will always take this extra time to change between the edge and
remote energies during MAD experiments.
Despite the Pilatus
6M relatively large pixel size compared to CCD detectors or the Eiger, in our
experience it is possible to resolve closely spaced diffraction spots
(see for example, the PDB structure 3M8C); note that, unlike the CCD,
this type of detector has a zero point spread function, which
contributes to limit the spot size.
The Pilatus 6M and Eiger 16M have a large dynamic range: 1,048,576 counts. When
the dynamic range is exceeded, the counter starts from zero again. The
only indication for this is that holes may be observed in
peaks. Blu-Ice marks in yellow pixels above 64,000 counts. These are not
overloaded, but the coloring makes it easier to spot true overloads.
Unlike an integrating detector the pixel array detectors have a
count-rate limit (the counts per unit of time rather than the total
number of counts over the entire exposure). Pixels that exceed the
maximum count rate (recorded in the image header) are flagged in red.
If in doubt, use the default values: in the Blu-Ice Collect tab,
click the Default button; in the Screening Tab, click Reset
defaults. If your crystals diffract very poorly or you are collecting
the images at extra long or short wavelengths you may have to increase
the time.
For radiation damage sensitive
samples, the best strategy is to do a two-wavelength MAD or SAD
experiment without exceeding the maximum recommended dose . Overdosing the crystal
results in a unit cell expansion which most often prevents accurate
measurement of any kind of phasing signal in the data. Radiation
induced intensity difference are no easier to measure than anomalous
or dispersive differences in this case.
For some derivatives (e.g., brominated DNA), the heavy atom may
become cleaved at very low doses. The program SHARP has been reported
to deal well with this particular case, using the loss of occupancy of
the anomalous scatterer to enhance MAD or SAD phases, as long as the
total dose is kept to a reasonable value .
BL12-1 and BL12-2
BL12-1 and BL12-2 are an undulator beamlines with microfocus capabilities and high
flux. Here are the major characteristics:
- Variable beam size : While other SSRL beamlines
operate at a fixed focus, and the beam size is changed by opening or
closing collimating slits upstream of the sample. On BL12-1, however,
the beam can be focused down to 40x5 (h/v) microns and on BL12-2 down to 40x15 (h/v) microns continuously.
Important: Because the full beamline flux can be concentrated on a
very small spot, it is important to avoid overexposing the
samples.
In addition, on BL12-2 it is possible to use smaller beam sizes (eg, 15x15) by inserting a microcollimator in the beam. To
use this feature, check the ''microbeam'' button of the
required size in Blu-Ice.
As a rule, the microbeam should only be used to study very
small crystals; for crystals larger than 10-20 microns it will be usually more
advantageous to use the ''normal'' beam focused to match the crystal
size.
- High zoom on-axis camera: Besides the sample camera
viewing the crystals from a below, a high zoom camera can be used to
see the sample from the beam direction. The on-axis camera video feed
can be accessed from the Blu-Ice video widget, by clicking the On-Axis
button. The Back Light can be inserted in conjunction with the
Light intensity bar to obtain the optimal sample
visualization.
- Automated beam optimizations using a special sample:The beamline software performs an automated
sample optimization periodically, using a special metal sample mounted
by the robot. This will take place between sample mounting and
dismounting cycles - ie, the software will wait until the sample
currently in used has been screened or collected from and dismounted
before mounting the special sample. The optimization takes about 10
minutes. After it is completed, the new crystal sample will be mounted
automatically. Since 2016, it is possible to stop the beam
optimization at any time. Doing this is not recommended if you
are using or planning to use the microcollimators, since they require
a very precise degree of component alignment. If using the
microcollimators, after skipping one or more optimizations, please
check the ``Optimize beam'' button in the hutch tab. If the color has
changed to green, use it to optimize the beam. Never use the ``Abort''
button to stop the optimization.
- Fast energy changes. Beam optimizations are not required
after a change of energy, so this only takes a few seconds. The only
exception is when changing the energy below or above 9keV, because the
gain of the Pixel Array Detector is energy-dependent and it needs to be
changed at that value; this procedure is automatic, and it takes about
2 minutes.
BL9-2
BL9-2 is a wiggler beamline. The beam size can be continuously changed by collimating the beam by a pair of slits. The slit size
is controlled by specifying the beam size in the Blu-Ice interface
- note that making the beam size smaller or larger than the limits
displayed in Blu-Ice will not change the beam size.
BL9-2 is equipped with a microspectrophotometer, useful to
monitor photoreduction when working with samples with a metal
center. See the Blu-Ice
documentation
for more information about this instrument.
Because beam optimizations are not required following a change of
energy, this is a very fast procedure. The only
exception is when changing the energy below or above 9keV, because the
gain of the Pixel Array Detector is energy-dependent and it needs to be
changed at that value; this procedure is automatic, and it takes about
2 minutes.
BL14-1
BL14-1 is a bending magnet beamline. The beam size can be
changed by collimating the beam by a pair of slits. The slit size
is controlled by specifying the beam size in the Blu-Ice interface.
SPEAR3 operation
The beamline status and current are displayed in
the Blu-Ice status
bar.
Additional information about SPEAR3 (including status of all
beamlines and the 24-hour fill history) is displayed on a monitor at
the beamline (top left corner of the console) and on the
SSRL SPEAR3 Web
page. See
also ''Monitoring the SPEAR3 status
remotely''
SPEAR3 operates in frequent fill mode. The
beam is topped up every 5 minutes. Data collection can
continue normally during the injection in frequent fill mode. The
normal variation of the beam intensity between fills is less than
1%. In this mode, the displayed current will always be close
to the maximum injected current.
Under normal data collection conditions, we cannot detect any substantial differences between the quality of the
data collected in frequent fill mode and with constant stored
beam. It is also impossible to determine whether an injection took
place during the collection of a diffraction
image by examining the reflections in that image.
If a manual injection takes place, the data collection software
will stop and restart when the temperature of the beamline optics
has stabilized. If an image was being collected during manual
injection, it will be recollected.
A message is
displayed in the Blu-Ice status box while the beam is stabilizing
following a manual injection.
Completely search the hutch for persons before activating the search reset.
The hutch door must be closed and locked before the search alarm
stops ringing, otherwise the procedure must be repeated.
Problems during data collection
If the beamline is open and Blu-Ice repeatedly displays the
message ''waiting for beam'', check that the beamline stoppers switch is
open on the key panel in the control rack (all green LEDs should be
lit). Remote users can see the stoppers LEDs by selecting the
appropriate panel video
preset
in Blu-Ice. If this is the problem, on-site users should
repeat the hutch search. Remote users must call staff or, outside
normal working hours, the duty operator (650-926-4040).
If the stoppers are open, try reoptimizing the beam.
If the beam optimization does not solve the problem, call support
staff.
To determine the cause of blank diffraction images, follow these steps:
- If there is no image displayed, try opening the image with a
different program (e.g, ADXV) If the image looks only blank on
Blu-Ice, the image server may have crashed. Contact support
staff (please, send an e-mail during non-working hours, as this
problem does not affect data collection).
- Check the contrast in the image
display: Images with no diffraction spots displayed at a high
contrast level can hide diffraction features for thin-sliced images.
- Verify that there is beam in the hutch (see the previous
question). Try reoptimizing the beam.
- Check the beam attenuation: The attenuation level depends on the
beam energy, so a filter combination appropriate for data collection at a high
energy can fully block the beam if you have changed to a lower
energy.
- Check that the detector cover is not on. Remote users can use
the ``Overview'' preset in the hutch video
in Blu-Ice.
- If nothing is blocking the detector look at the shutter
controller to determine if the shutter is opening. The switch on
the controller should be on "auto" and a red LED light should light
up when the shutter is open, as shown in
shutter-controller. Remote
users can use the Panel
camera
to view the shutter controller. In addition, when the shutter opens
you should be able to see an increased reading for the
I_beamstop beam
monitor.
The I_beamstop reading will be low at long wavelengths (low
energies). Verify that there is beam on the beamstop by going to a
shorter wavelength (higher energy).
Figure 41:
Shutter controller showing open shutter status
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- If there is diffuse diffraction but no spots, check the
centering of the crystal. Open up the slits to increase the beam
size and collect images at different crystal orientations. If you
get diffraction with a large beam size or at some phi positions but
not others, the phi axis may be misaligned. Contact support staff.
- If there is no difference with large slits or at different phi
positions, the crystal may not diffract (even if it was diffracting
previously). Check the cryojet temperature (Blu-Ice will display an
error in the status display if the temperature raises above 120 K);
dismount the sample and try another sample. If you do not observe any
diffraction or scatter from any sample, contact support staff.
The first time you log in to Blu-Ice it will request your
password. Make sure that it is typed correctly. If you cannot log in,
contact the user support staff and let them know what Unix account you
are using.
If you cannot start Blu-Ice from the icon in the XFCE
menu:
- Open a terminal on the local workstation or NX client.
- Log in to a different beamline workstation. For example, if you
are trying to open Blu-Ice from bl92a, log in to bl92b or bl92c. Use
the command:
>ssh bl92a
>go
Note: The
NX client tries to open Blu-Ice on the ''c'' workstation; if Blu-Ice
fails to open, make sure that you use the ''a'' or ''b'' computer.
- Verify that the Blu-Ice window is active.
- If you lose permission to connect to Blu-Ice (the DCSS server is off), contact support staff.
- If some buttons are inactive (grayed out) move the mouse over the
button. Blu-Ice will display a message explaining
why the button is inactive. If the message tells that the
control software for a hardware component is off-line (''DHS
off-line'') contact support staff.
- If the Blu-Ice status window displays the messages ''MOTOR STOP
BUTTON LATCHED'', an emergency stop button may have been depressed
accidentally. To reset the motors, press the green motor
reset button. If collecting data remotely call support
staff, or, after working hours, try contacting the duty operator at
650 926 4040.
- If the entire Blu-Ice interface fails to respond, there might
be a pop-up window showing an error hiding behind another window. If
you cannot find it or cannot clear the error, try exiting the Blu-Ice client and
starting a new one. If the fault persists, call support staff.
- Sometimes, a computer crash or a network problem can hang the system. In
this case, other programs and processes will also be affected. If the
window manager program is hanging, log on to the host computer from
another terminal and kill the processes. To list processes:
>ps -u "your_id"
and kill them with
>kill -9 "process_id"
- If all the computers at the beamline are hanging or data
collection will not proceed, the file system may have crashed. Contact
support staff.
A detector error message in Blu-Ice can have many different
causes; although often staff intervention is required in order to
continue data collection, it is a good
idea to retry the image collection before calling staff, as
some errors (e.g., a transient network glitch) do not disable the
detector permanently.
Important: Note that the ''Detector Error'' message will still be displayed
after a problem with the detector has been fixed; the message will only
disappear once an image has been collected without errors.
It is advised to optimize:
- After changing the wavelength from the Hutch tab.
Automatic optimization is performed after wavelength
changes during MAD data collection.
- After changing the beam size, specially when going from the
large to a smaller beam size, or on BL12-2, when using a
microcollimator several hours after the last optimization.
Automatic optimization is
also performed at regular intervals during data collection and
therefore, manual optimization should not be necessary.
Most crystallographic software packages are only installed on the
data processing servers and not on the local beamline
computers. Graphics programs are installed only on the local beamline workstations.
Check the relative load of the data processing server as described in
''data processing environment''.
Why can't I autoindex my images?
The detector may have been
offset from Blu-Ice: Check the detector positioner vertical and horizontal values
in the Blu-Ice hutch tab. If they are not 0, look at the center
coordinates in the image header.
The program LABELIT, and the mosflm
and autoxds scripts
provided at the
SSRL can use the image header coordinates directly for
autoindexing. However, if you use HKL3000, iMosflm or have your own
scripts to run XDS, you must specifically provide the offset center as
follows:
- For HKL3000 or iMosflm:
y = Detector height (mm) - Cy (CCD detectors and Pilatus)
y = Cy (Eiger)
x = Cx (all detectors)
- For XDS:
x = Cx/pixel size (all detectors)
y = Detector height (pixels) - Cy/pixel size (CCD detectors and Pilatus)
y = Cy/pixel size (Eiger)
Indexing may also fail if the diffraction is weak,
if there are many ice rings or if there is a double lattice. Editing
the spots manually often circumvents these
problems.
Local users can mounting samples manually determine the accurate
center position by following these steps:
- Move the detector to the distance used for data collection.
- If the resolution at the edge of the detector is 3Å or higher,
use the Si sample, otherwise use the polyethylene sample. Both
samples are located in the same compartment on the beamline tool board.
- Collect a diffraction image. For the Si sample, use a
delta phi of 15 degrees and 5 s exposure time.
For the polyethylene sample, use a delta phi of 0.02 degrees and a 1 s
exposure time. If the detector saturates, attenuate the beam.
- Run the program center on one of the pxproc servers
to calculate the direct beam position from the image.
Important: The beam center position in the image header should be accurate to within 0.1mm. In
the extremely rare event that it is off my a larger amount, make
sure to tell user support staff.
You can use the screening tab
to
display the crystal snapshots.
You may also use the program display (from a Linux or Unix shell).
On-line Space Group diagrams are available at
http://smb.slac.stanford.edu/facilities/software/spacegroups/. This site is for
local access only; the NX Client can be
used to view the tables remotely.
The full International Tables are available at
http://it.iucr.org/. The IUCr
site is fully accessible from SSRL or the NX client (or off-site if your local institution
has a license).
Users are responsible for backing up by their data by the end
of their beam time. Images stored on the /data disk can be deleted at any time.
Special requests to keep files on /data should be made
to support staff. The /home area can be used to store small
files indefinitely.
Remote access
How can I monitor the SPEAR3 beam remotely?
The beamline status and current SPEAR3 intensity are displayed in
Blu-Ice. The video tools in Blu-Ice can also be
used to look at the beamline SPEAR
monitor;
in addition, the SPEAR3 status and fill history can also be accessed
via the
web.
For updates or inquiries about SPEAR3 you can call the duty
operator 24/7 at 650 926-4040, or the beam information line at
650 926-BEAM (2326).
HKL3000 displays a screen size error
In order to run HKL3000, the NX client window must
be at least 1100 x 900 pixels. If HKL3000 gives the error ''HKL2000
requires screen width larger then 1100 and screen height larger then
900 (sic)'', enlarge the NX client window as described in the remote desktop configuration
(see section "Tuning the configuration" near the end of the page).
There is a limit on the number of ADXV processes allowed to run
on the NX server in order to stop it from running out of memory
(currently the limit is 5). To inspect new image, close some old ADXV
windows.
Please consult the remote desktop documentation.
Click on the Settings icon in the Xfce
panel to access the Settings Manager GUI.
Exception: Use only the Blank Screen as a screen saver (animated
screen savers use a lot of CPU, which can affect remote access through
the NX client).
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![\includegraphics[width=0.5\textwidth]{images/shuttercontrol.ps}](img26.png)