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analySIS®

MegaViewIII
Installation
Guide
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Olympus Soft Imaging Solutions GmbH, Johann-Krane-Weg 39, D-48149 Münster, Tel. (+49)251/79800-0, Fax: (+49)251/79800-6060
About this document
This document describes the installation procedure for the Mega-
ViewIII system on to a generic TEM column. It includes the following:
the installation of hardware components on the PC running
analySIS ®; the MegaViewIII camera unit on to the wide angle port of
the microscope; the pneumatic components required for controlling
the camera position; cabling; proper set up of the camera for regular
use; some handling and usage precautions; some servicing infor-
mation; and some technical data.
Please read these instructions before attempting an installation.
Some parts or procedures may differ for different models of micro-
scope, but it should not cause any major problems.
MegaViewIII
Contents

MVIII Installation Guide


Quick-start Installation 2

Handling and Usage Precautions 3

The MegaViewIII Camera Unit ............................................3


The FireWire board IEEE-1394 ...........................................4
The MegaViewIII Controller .................................................4

Camera Installation Procedure 5

FireWire board IEE-1394 .....................................................5


MegaViewIII unit ..................................................................6
MegaViewIII controller unit ..................................................8
Cables ................................................................................10

Software Installation under Windows 2000 12

Remote control software ....................................................12


analySIS, camera driver, and remote control ....................12
Camera cable ....................................................................13
Troubleshooting .................................................................15

Software Installation under Windows XP 21


HWMVIII0602

Remote control software ....................................................21


analySIS, camera driver, and remote control ....................21
Camera cable ....................................................................22
Camera driver installation ..................................................23
MegaViewIII • Contents

Camera set-up under analySIS 28

Selecting an input channel ................................................ 28


Acquiring images ............................................................... 29
Defining gain and offset correction .................................... 29
Calibrating input channel ................................................... 37
Activating the remote control ............................................. 39
Magnification setting .......................................................... 40

Servicing Notes 42

Adjusting the MegaViewIII In/Out movement speed ......... 42


Adjusting the prism position .............................................. 43
Focusing the camera ......................................................... 44
Focusing the camera with beam stop ................................ 46

Appendix 48

Technical specification ...................................................... 48


Packing List ....................................................................... 48

CE DECLARATION OF CONFORMITY 49
MegaViewIII • Quick-start Installation

Quick-start Installation
This section serves as a guide for those who have installed the MegaViewIII
system before. It is a quick reminder for a sequence of steps that should be
followed for a successful installation.
If you are installing for the first time or you are unsure of the procedure then
please skip this section and read all other sections carefully before
beginning the actual installation.
The basic installation steps are:
1) Install a FireWire TM board if not yet existing.
2) Evacuate the TEM column.
3) Remove TEM flange cover.
4) Remove protective cover on prism.
5) Install MegaViewIII unit.
6) Connect pneumatic tube with T-joint to pressurized air supply.
7) Connect pneumatic tube from MegaViewIII controller unit (connector
Air In) to self-closing valve.
8) Connect pneumatic tubes from MegaViewIII controller (connectors A
and B) to MegaViewIII camera unit (the tube from A belongs to the
connector closer to the prism).
9) Connect MegaViewIII In/Out position control box (handbox) to the
MegaViewIII controller (connector Handbox).
10) Connect the cable from the MegaViewIII controller (connector
Computer) to the serial port on PC (normally COM2).
11) Connect the Remote Control cable from TEM to PC serial port
(normally COM1), if this option is available.
12) Install Remote Control software, if this option is available. Note that for
some microscopes, the remote is installed during installation of
analySIS ®.
13) Install analySIS ® software.
14) Install camera driver (Windows XP only).
15) Connect the cable from the MegaViewIII camera unit to the FireWireTM
board on PC
2

16) Set-up remote control for all input channels, if this option is available.
17) Focus the camera.
18) Set-up offset and gain correction image for the two image formats.
19) Calibrate input channels.

related topics
FireWire board IEE-1394 p. 5
MegaViewIII unit p. 6
Software Installation under Windows 2000 p. 12
Software Installation under Windows XP p. 21
Camera set-up under analySIS p. 28
Focusing the camera p. 44
Handling and Usage Precautions

Handling and Usage Precautions


Important Please read the following carefully before attempting installation of
camera.

The MegaViewIII Camera Unit


Warning! • Install analySIS ® first before connecting the camera to the
- Windows 2000 FireWire TM board.

General Warnings! • The MegaViewIII is shipped fully assembled, tested and ready for
mounting onto the microscope. Do not adjust any part within this
assembly unless focusing is required.
• The quality of the image should be evaluated to decide whether
focusing of the MegaViewIII lens system is required. Instructions to
perform this task can be found in the ”Servicing Notes” section.
Fully assembled
MegaViewIII camera unit
ready for shipping.

3
• The prism is shipped with a protective cover mounted over it. The
fluorescent coating on the top surface of the prism is very delicate and
is easily damaged. Remove this cover just before mounting the unit
onto the microscope. Do not touch the fluorescent coating with
anything. You may clean it by gently blowing clean dry air over it.
MegaViewIII • Handling and Usage Precautions

Prism with cover installed.


Remove the two small
screws to remove the cover
just prior to mounting onto
the microscope.

• Make sure that the camera-prism mechanism is in the OUT position


before installing.
• Do not open the throttle valves (which controls the speed of movement
of the camera unit) too large. This may cause the unit to move with too
much force and cause mechanical damage.
• Exposure to high temperature or humidity will affect the characteristics.
Avoid storage or usage in such conditions.
• Do not subject the camera to too much mechanical shock.
• The FireWireTM cable plug is not symmetrical. You must make sure
that the cable plug is facing the same way as the camera socket when
you plug the cable into the camera connector.
• Do not disconnect the FireWire TM cable while analySIS® is running.
• Wear protective glove when you handle the camera.

The FireWire board IEEE-1394


Warnings! • The components of these devices are easily damaged by static
discharge. Wear grounding wrist straps when handling them.
• Do not plug in or unplug the cards from the PCI slot while the PC is on.
4

• Please follow the cable installation procedure in this booklet step by


step to avoid any incorrect connections. Incorrect connections may
result in serious damage to your PC system and/or the camera.

The MegaViewIII Controller


• If for any reason that you need to disconnect the pressurized air supply,
make sure that the camera-prism mechanism is in the out position.
You may encounter some problems in moving the camera-prism
mechanism later if pressurized air is reconnected when it is in the in
position.
• The air pressure requirement is 4 - 6 bars. Air pressure outside this
range may cause the unit to operate incorrectly or cause mechanical
damage.
Camera Installation Procedure

Camera Installation Procedure


FireWire board IEEE-1394
when is this necessary? This procedure is necessary only if the PC does not have a FireWireTM on-
board hardware configuration or if it is not yet equipped with a FireWireTM
board.
the FireWire TM board

How to install the FireWire board


1) Turn off the PC and disconnect the power supply.
2) Remove the cover from the PC.
3) One PCI slot is required for the FireWire TM board. Insert the board into
the slot.
FireWireTM board installed

4) Replace the cover onto the PC.


MegaViewIII • Camera Installation Procedure

5) You are now ready to first install the camera onto your microscope,
then to install analySIS® onto your computer, and last to connect the
camera with your conputer.

MegaViewIII unit
How to install the MegaViewIII
1) Remove the elliptical housing. There are two small screws located at
the bottom of the housing.
Note You are normally not required to disassemble any part of the MegaViewIII
system (besides the housing) upon installation except for the purpose of
focusing the lens system.
Remove the two housing
screws and remove the
camera housing.

2) Check the distance from front of the flange surface to the middle of the
prism. This distance varies with the model of the microscope.
Note The camera is delivered having the prism position preadjusted to your type
of microscope. So usually, you don’t have to change the prism position. If
6

you have to do so please follow the servicing notes.

3) Check that the prism holding lugs and the prism rod holding lugs
are sufficiently tight.

related topics
Focusing the camera p. 44
Adjusting the prism position p. 43
MegaViewIII unit

Checking prism holding lugs (left) and prism rod holding lugs (right).

4) Vent the camera chamber of the microscope, then remove the camera
port cover flange.
5) Check that the O-ring and flange surface is clean.
Clean the O-ring.

7
6) Remove the two small screws holding the prism cover and remove this
protective cover. Check that there are no dust particles on the
fluorescent coating.
7) Place the MegaViewIII unit onto the camera port and tighten the
screws. The MegaViewIII unit is rather heavy and may require one
person to hold it in place while another tightens the screws. It may be
necessary to use straight hex ball drivers to tighten the screws.
Regular L shaped Allen-key drivers may not be practical due to space
constraints around the flange screws. This situation may vary from
model to model of TEMS.

related topics
General Warnings! p. 3
MegaViewIII • Camera Installation Procedure

Tighten the screws that hold


the MegaViewIII unit on to
the microscope. Note that
the flange and position of
the screws may differ from
microscope to microscope.

8) Mount the elliptical housing on the MegaViewIII unit.

MegaViewIII controller unit


4 pneumatic tubes The MegaViewIII controller unit comes with 4 pneumatic tubes.
1) One tube with a T-join on one end and a self-closing valve on the other.
2) One tube with a connector to the self-closing valve on one end.
3) 2 bare tubes.
Pneumatic tube with a T-join
on one end and a self-
closing valve on the other Self-closing valve
end. Note that the size and
type of pneumatic fittings
may differ depending on the
model of the microscope.

T-join
8
MegaViewIII controller unit

The tube with the connector


to the self-closing valve
should be connected to the
IN inlet of the MegaViewIII Self-closing valve
controller unit.

How to connect the tubes and the controller box cables


1) Locate the pressurized air system and a pneumatic tube with air
pressure between 4 and 6 bars. Turn off the air pressure and cut this
tube, then connect the T-join in between. This tube may also be
connected directly to a free pressurized air outlet – in this case the T-
join will not be required. The open ends of the tubes are connected as
shown in the figure.

Connecting an open ended


pneumatic tube to an air
inlet/outlet.

2) The second tube with the connector to the self-closing valve on one
end should be connected to the inlet marked Air In on the front of the
MegaViewIII controller unit.

9
The back panel of the
MegaViewIII controller box,
showing the serial
connectors to the PC and
the Handbox, and the inlet
and outlets for pressurized
air.

3) The other two bare tubes should be connected to the outlets marked A
and B on the MegaViewIII controller unit. These tubes should be
connected to the correct inlets on the MegaViewIII unit: The tube
connected to outlet A on the controller unit side has to be connected to
the inlet which is closer to the prism on the camera side.
MegaViewIII • Camera Installation Procedure

Pneumatic Inlets on the


MegaViewIII unit.

4) Connect the self-closing valve to the corresponding connector by just


pushing it until it clicks into place. It can be removed by just firmly
pulling back the spring-loaded release of the self-closing valve.

Cables
1) Don’t connect the camera with the computer at this step - before having
installed analySIS®.
2) Connect the MegaViewIII In/Out position control box (handbox) to the
serial port marked Handbox on the MegaViewIII controller box using
one of the DB9-DB9 serial cables. This box is used to manually move
the prism between the IN and OUT positions.
The MegaViewIII In/Out
position control box
(handbox). This box must
be connected to the serial
port marked Handbox on
the MegaViewIII controller
box.
10

3) Connect the MegaViewIII controller box with a free serial port of your
PC (normally COM2) using the other DB9-DB9 serial cable. Tighten the
connectors with a small screwdriver.
4) Some microscopes have a remote control feature. This may require an
additional serial cable connection between the microscope and the
analySIS ® PC (typically COM1).
Cables

11

The diagram shows a typical installation on a TEM. Microscopes may have the camera port at different locations.
MegaViewIII • Software Installation under Windows 2000

Software Installation under Windows 2000


Remote control software
Remote Control driver Remote control is available for some later models of microscopes. It allows
software analySIS® to read some microscope parameters, such as magnification, via
the serial port. It is also used to send control commands to the microscope,
such as moving the sample stage.
Certain models of microscope require a separate installation of the Remote
Control driver software. This should be done before installation of
analySIS®. This software is normally supplied on a seperate disk with
installation instructions.
Some models of microscope will need the remote drivers to be installed
during installation of analySIS®.

analySIS, camera driver, and remote control


Warning! analySIS® must be installed first before connecting the camera to the
FireWireTM board.

system requirements The MegaViewIII camera requires analySIS® (3.2 version), and Microsoft
Windows 2000 (or later). analySIS ® also supports multithreading on
multiprocessors, thus making full use of the power of the latest multi-CPU
PC’s.
camera driver installation Installation of the MegaViewIII driver is simply done via the analySIS®
setup. You are only required to select the FireWireTM board and specify the
additional camera driver during setup.
analySIS® setup Installing analySIS® is a simple process following on screen instructions.
Follow analySIS® setup
wizard for FireWire board
selection and camera driver
installation. Select the
Firewire checkbox in the
12

Input Device (Digitizer)


Selection dialog box.
analySIS, camera driver, and remote control

Then select the Soft


Imaging System
Manufacturer entry and the
MegaView III Camera entry.

Note analySIS ® can support several different types of FireWireTM cameras


simultaneously.
Select the appropriate
model of microscope, if
remote and respective
driver are available.

In the Stage Selection


dialog box select the
appropriate stage control, if
remote and respective

13
driver are available.
MegaViewIII • Software Installation under Windows 2000

Camera cable
Warning! analySIS® must be installed first before connecting the camera to the
FireWireTM board.

How to connect camera and computer


1) Plug one end of the FireWireTM cable into the camera connector.
" The FireWireTM cable plug is not symmetrical. You must make
sure that the cable plug is facing the same way as the camera
socket when you plug the cable into the camera connector.
The camera chip may be ruined otherwise.
Make sure the cable plug is
facing the right way before
you plug it into the camera
socket.

2) Plug the other end into one of the small connector sockets on the
FireWireTM board.
" There are two or more connector sockets depending on the
FireWireTM board. You can use any of them.
" If the Digital Signature Not Found MS-Windows dialog box(es)
appears on your screen. Confirm the message as often as
necessary until the dialog boxes disappear.
14
Troubleshooting

3) Open the Windows Device Manager via Start > Settings > Control
Panel > System > Hardware > Device Manager....
" The Device Manager dialog box contains the correct MegaViewIII
driver entry SoftHard Technology MV4.0 Camera. The camera is
ready for use.

Windows Device Manager shows the IEEE 1394 FireWireTM board and the
MegaViewIII driver SoftHard Technology MV4.0 Camera.

Troubleshooting
what to do when the When the MegaViewIII camera driver has been installed by analySIS®
camera has been before connecting the camera to the computer, Windows 2000 will correctly

15
connected before recognize and use the installed driver. Yet if the camera has been plugged
analySIS ® installation into the computer before the MegaViewIII camera driver installation by
analySIS ®, Windows 2000 installs its own standard FireWireTM camera
driver, meaning that the MegaViewIII camera can’t communicate with the
computer. Installing analySIS® now doesn’t install the correct camera driver
since this is forbidden by Windows 2000. Here, you have to install the
correct camera driver from the software’s setup CD separatly from the
software installation. Do the following:
1) The camera has been connected to the computer before installation of
analySIS ®. Then the Found New Hardware Wizard MS Windows
dialog box will appear at that moment when you connect camera and
computer.
MegaViewIII • Software Installation under Windows 2000

2) Click on the Next button.


16

3) Select the Search for a suitable driver for my device (recommended)


option and click on the Next button.
4) Insert the analySIS ® setup CD into your computer’s CD-ROM drive.
Minimize the appearing setup dialog box.
Troubleshooting

5) Make sure that only the CD-ROM drives check box is selected and click
on the Next button.
" Wait until Windows has found the correct camera driver.

17

6) Click on the Next button.


" The Digital Signature Not Found dialog box appears on your
screen.
MegaViewIII • Software Installation under Windows 2000

7) Click Yes.
" Wait until the camera driver has been installed.
18

8) Click on the Finish button.


" The Digital Signature Not Found MS Windows dialog box(es)
appear on your screen. Confirm the message as often as
necessary until the dialog boxes disappear.
Troubleshooting

9) Open the Windows Device Manager via Start > Settings > Control
Panel > System > Hardware > Device Manager.... The Device Manager
dialog box contains the correct MegaViewIII camera driver entry:
SoftHard Technology MV4.0 Camera.

19

Windows Device Manager shows the IEEE 1394 FireWireTM board and the
MegaView III camera driver SoftHard Technology MV4.0 Camera.

10) Install analySIS®. During the installation check the Firewire check box
to in the Input Device (Digitizer) Selection dialog box. Select the
MegaViewIII check box in the following Camera Selection dialog box.
11) Shut down the computer and restart it.
MegaViewIII • Software Installation under Windows 2000

" The Digital Signature Not Found MS Windows dialog box(es)


appear on your screen. Confirm the message as often as
necessary until the dialog boxes disappear.

12) Open the Windows Device Manager via Start > Settings > Control
Panel > System > Hardware > Device Manager.... The Device Manager
dialog box contains the correct MegaViewIII camera driver entry:
SoftHard Technology MV4.0 Camera.
20

Windows Device Manager shows the IEEE 1394 FireWireTM board and the
MegaViewIII camera driver SoftHard Technology MV4.0 Camera.
Software Installation under Windows XP

Software Installation under Windows XP


Remote control software
Remote Control driver Remote control is available for some later models of microscopes. It allows
software analySIS ® to read some microscope parameters, such as magnification, via
the serial port. It is also used to send control commands to the microscope,
such as moving the sample stage.
Certain models of microscope require a separate installation of the Remote
Control driver software. This should be done before installation of
analySIS ®. This software is normally supplied in a separate disk media and
installation instruction.
Some models of microscope will need the remote drivers to be installed
during installation of analySIS®.

analySIS, camera driver, and remote control


system requirements The MegaViewIII camera requires the 3.2 analySIS ® and Microsoft
Windows XP (or higher). analySIS® also supports multithreading on
multiprocessors, thus making full use of the power of the latest multi-CPU
PC’s.
camera driver installation Installation of the MegaViewIII camera driver starts via analySIS ® setup.
You are required to select FireWireTM and specify the additional camera
driver during setup. Then you have to get Windows to use the MegaViewIII
camera driver.
analySIS ® setup Installing analySIS® is a simple process following on screen instructions.
Follow the analySIS® setup
wizard for FireWire selection
and camera driver
installation. Select the
Firewire checkbox in the
Input Device (Digitizer)
Selection dialog box.

21

related topics
Camera driver installation p. 23
MegaViewIII • Software Installation under Windows XP

Then select the Soft


Imaging System
Manufacturer entry and the
MegaView III camera entry.

Note analySIS® can support several different types of FireWireTM cameras


simultaneously.
Select the appropriate
model of microscope, if
remote and respective
driver are available.

In the Stage Selection


dialog box select the
appropriate stage control, if
22

remote and respective


driver are available.
Camera cable

Camera cable
How to connect camera and computer
1) Plug one end of the FireWire TM cable into the camera connector.
" The FireWireTM cable plug is not symmetrical. You must make
sure that the cable plug is facing the same way as the camera
socket when you plug the cable into the camera connector.
The camera chip may be ruined otherwise.
Make sure the cable plug is
facing the right way before
you plug it into the camera
socket.

2) Plug the other end into one of the small connector sockets on the
FireWireTM board.
" There are two or more connector sockets depending on the
FireWireTM board. You can use any of them.
3) Continue with the camera driver installation before starting analySIS ®.

23
MegaViewIII • Software Installation under Windows XP

Camera driver installation


The MegaViewIII camera driver is not signed and therefore not used
automatically under Windows XP. After having installed the analySIS® and
connected the camera to the computer - or vice versa - you have to go
through the following steps to get Windows XP to use the SIS View camera
driver:
1) Open the Windows Device Manager via Start > My Computer > (right
mouse) > Properties > Hardware > Device Manager....

" The Device Manager dialog box shows the Generic 1394 Desktop
Camera entry in the Imaging devices section. The exclamation
mark indicates a device conflict.
24

2) Select the Generic 1394 Desktop Camera entry and click on the
Update button in the device manager button bar.
Camera driver installation

3) Select the Install from a list or specific location (Advanced) option and
click on the Next button.

25

4) Select the Don’t search. I will choose the driver to install. option and
click on the Next button.
MegaViewIII • Software Installation under Windows XP

" The Model list contains the SoftHard Technology MV4.0 Camera
entry.
5) Click on the Next button.
" The next Hardware Update Wizard dialog box will be opened in the
background displaying the message to wait while the Wizard
installs the software.
" Additionally, the Hardware Installation dialog box will pop up in the
foreground.
26

6) Click on the Continue Anyway button.


" The Hardware Update Wizard dialog box which had been in the
background, will become visible.
Camera driver installation

" After having finished the MegaViewIII camera driver installation the
following dialog box will be opened.

27

7) Click on the Finish button to end the hardware update wizard.


" In the Device Manager dialog box there is still the same Generic
1394 Desktop Camera entry but the exclamation mark indicating
the device conflict should have disappeared. The camera is now
ready for use.
MegaViewIII • Software Installation under Windows XP

" If the exclamation mark is still there it may be worthwhile to repeat


the whole procedure once - beginning with the analySIS ® setup.
28
Camera set-up under analySIS

Camera set-up under analySIS


Selecting an input channel
How to select an input channel
1) Switch on your TEM and the MegaViewIII Controller Box.
2) Move the camera into the TEM column.
3) Start analySIS®.
" The WaaCtrl dialog box will be opened, if the controller box is not
correctly connected.

4) Check the connection and then select the respective Com-Port, usually
COM2. Click OK.
5) Select the Image > Set Input command.
" The Set Input dialog box will be opened. It contains three pre-
defined input channels for your MegaViewIII camera: MegaView III
Search, MegaView III Focus and MegaView III Snapshot.
You can select an input
channel out of three
predefined channels.

29

Channel overview
Settings Search Focus Snapshot
Resolution 688 x 516 688 x 516 1376 x 1032
Binning 2x2 2x2 1x1
Exposure time 50 ms 50 ms 200 ms
Automatic gain display on on on
Online histogram on off on
Real time FFT off on off
Shading correction on off on
MegaViewIII • Camera set-up under analySIS

MegaView III Search " MegaView III Search is for locating relevant parts of a specimen
quickly and easily. In this case, high resolution is of lesser
importance than a rapid frame rate.
MegaView III Focus " MegaView III Focus is a channel with which you can interactively
adjust and focus your TEM. The real-time Fast Fourier
Transformation (RT-FFT) will aid you in doing so.
MegaView III Snapshot " MegaView III Snapshot is a channel for actually acquiring images.
This channel has the highest resolution available.
6) Select, e.g., the Mega View III Search channel.

Acquiring images
1) Select the Mega View III Search channel.

2) Click on the Standard (button bar) > Acquire button.


3) The camera’s live image will be displayed in the image window.
" The online histogram window will be opened. The diagram shows
the current gray-value distribution of the live image.
Online histogram.

4) Select the Image > Camera Control... command.


" The Camera Control dialog box will be opened.
" Change the exposure time, if necessary.
Camera Control.
30

5) Locate the desired sample position.


6) Click on the Standard (button bar) > Snapshot button.
" The live image will be frozen.

Defining gain and offset correction


Shading correction is required for the acquisition of best quality images. One
gain image and one offset image are required for each format used in the
input channels for the purpose of automatic shading correction. In fact, you
will mainly use the shading correction for your snapshot channel(s), i.e., at
high resolution, for shading correction reduces the speed in the live modus.
But you also need it when you focus your camera.
Defining gain and offset correction

Online shading The Online shading correction is how analySIS® corrects fixed image
correction structures (resulting from the imaging system) during acquisition.
What’s shading • Irregular illumination on the CCD chip can, eg, lead to a dip in intensity
correction for? at image borders. This means that image edges are darker than the
central image area, thus specimen structures and image gray values
are no longer in clear correlation with one another. Setting of
thresholds is impossible and successful automatic measurement
operations are also not feasible.
• The amplification of individual camera-array pixels can fluctuate slightly
in CCD cameras. Individual pixels - so-called "warm pixels" - are thus
somewhat brighter or darker than neighboring pixels which were
illuminated with the exact same intensity. This phenomenon can be
particularly disturbing when working with long exposure times.
• Luminescent screen thickness in the reflecting prism can vary slightly,
resulting in fluctuations in light yield.
What does shading The following image operations are executed during a real-time shading
correction involve? correction:
1) The offset image is subtracted from the original image.
2) The resulting image will then be divided by the gain image - (the gain
image has already been offset corrected at this point - i.e., the
appropriate offset image has already been subtracted from the gain
image).
3) The resulting corrected image will then be multiplied by the mean gray
value of the offset-corrected gain image. Mean image brightness is
thus not affected by shading correction.
Shading correction Prerequisite for shading correction is your having acquired a valid gain
prerequisites image and a valid offset image - using the Image > Acquire Reference
Images... command. When you begin image acquisition and analySIS®
cannot find any valid reference images, you’ll be instructed to acquire
reference images.
Offset image A camera offset image is acquired without any illumination. An offset
correction is necessary when using a CCD camera in order to compensate
for a non-homogenous dark-current signal. A shading correction involves

31
subtracting the offset image from the original image.
To be precise, two offset images with varying exposure times are acquired
and saved. Common exposure times are 50 ms and 1000 ms. A third offset
image - when necessary - is then computed via inter- or extrapolation of the
first two offset images for the current exposure-time setting. If you alter
exposure time, and then acquire an image using real-time shading
correction, the requisite offset image will be automatically recomputed -
without your noticing.
Gain image The camera gain image is acquired at homogeneous illumination - without
any specimen. The original image is divided by the gain image during
shading correction.
Both reference images will automatically be saved to the computer hard
disk.
Warnings! • If you have to work with more than one TEM acceleration voltage you
will have to acquire reference images for each of these. If analySIS®
can directly read out the voltage set on your TEM via remote,

related topics
Focusing the camera p. 44
MegaViewIII • Camera set-up under analySIS

analySIS ® will automatically select the correct reference images as


needed. If you do not have remote control you will have to acquire a
new gain image anytime you alter the voltage level.
• You will also need to have reference images for each image format
which is defined in an input channel and used to acquire images with.

How to acquire an offset image


1) Switch on your TEM and the MegaViewIII Controller Box.
2) Make sure that no light can reach the CCD chip of the camera. To do
this, cover up the window of the TEM projection chamber and turn off
the electron beam. Move the camera into the TEM column.
3) Select the logical input you’re acquiring reference images for - e.g., the
high resolution Mega View III Snapshot channel.
4) Select the Image > Acquire Reference Images... command to acquire
the offset image for correction of camera dark current.
" The Acquire Reference Images dialog box will be opened.
Acquiring offset image.
32

5) Use the pre-set value of 4 in the Offset Image > Averaging cycles field.
" An averaging cycle of 4 means that 4 images will be read in
successively and the arithmetical mean will be saved as the
reference image. (To be exact, there are in fact eight images being
acquired - four images twice - at two different exposure times.)
" The control lamp will indicate whether there is an valid reference
image existent (green) - or there is not (red).
" The word missing will appear beside the control lamp if there is no
valid reference image available, or if there is one, that image’s
creation date.
6) Click on the Acquire button.
" You’ll be asked to switch off the illumination. Then press OK.
" The offset image will be acquired.
" The warning lamp will now be green - the date will be updated.
Defining gain and offset correction

A typical MegaViewIII offset


image (this image has been
contrast enhanced).
Any structures resulting
from a non-homogeneous
dark-current signal are
barely visible, but these can
- especially where low-
intensity acquisitions are
concerned - lead to artifacts,
which impede quantitative
image analysis.

7) Click on the Close button.

How to acquire a gain image


1) Make sure that illumination of the luminescent screen (on the topside
of the MegaViewIII reflecting prism) is even. Move the camera into the
TEM column, turn on the electron beam and remove the specimen from
the ray path.
" You can also make use of a hole through the specimen - instead of
removing the specimen - so that the beam can pass through it
unhindered.
" The electron beam intensity should be essentially the same as you
are acquiring images with.
2) Make sure, that the input channel features the online histogram:
Select the Image > Configure Input... command and select the Display
> Online histogram check box.

33
MegaViewIII • Camera set-up under analySIS

3) Select the Image > Camera Control... command.


" The Camera Control dialog box will be opened.
" Check the exposure time: It should be essentially the same as you
are acquiring your images with, i.e., it should be within a range of
about 20%.
Adjusting exposure time.

4) Select the Image > Acquire command to switch over to the live mode.
" A live image will be shown in the active Viewport.
" The online histogram window will be opened. The diagram shows
the current gray-value distribution of the live image.
5) Use the histogram to check whether the gray values are situated
centrally in the diagram. As necessary you can alter, e.g., electron
beam intensity or spot size of the electron microscope, and/or you
adjust the exposure time a little to shift gray values roughly into the
middle of the graph.
" The mean value shown ought to be between 45 % and 55 %.
The mean value should be
between 45 % and 55 %.

6) Freeze the image by pressing the Snapshot button.


34

7) Select the Image > Acquire Reference Images... command to now


acquire the gain image for correction of camera amplification.
" The Acquire Reference Images dialog box will be opened.
Defining gain and offset correction

Acquiring gain image

8) Use the pre-set value of 4 in the Gain Image > Averaging cycles field.

Acquiring gain image 9) Deselect the Automatic intensity adjustment check box.
without automatic 10) Click on the Acquire button.
intensity adjustment " You’ll be asked to remove the specimen and to check for
homogenous illumination. Do such and press OK.
" The online histogram window will be opened. The diagram shows
the current gray-value distribution of the live image.
" You’ll be asked to adjust the mean intensity to approximately 50 %.
11) Check, if the mean value is between 45 % and 55 %. Then press OK.
Otherwise, use electron beam intensity and/or exposure time to adjust
the gray value distributuion that way.
Then press OK.

35
Acquiring gain image 9) Select the Automatic intensity adjustment check box.
using automatic intensity 10) Click on the Acquire button.
adjustment " You’ll be asked to remove the specimen and to check for
homogenous illumination. Do such and press OK.
" The image analysis software will automatically set the optimum
exposure time. Using the optimum exposure time the online
histogram is centered.

" The gain image will be acquired and saved offset corrected. This
is why you can only acquire a gain image if a valid offset image is
available.
" The control lamp will now be green, and the date will be updated.
MegaViewIII • Camera set-up under analySIS

A typical MegaViewIII gain


image.
The non-homogeneous
illumination of the image
field is readily visible. The
image appears less
homogeneous than it in fact
is - this is because image
contrast has been
automatically enhanced for
monitor display.

11) Click on the Close button.


" All input channels that use a display format of 1376 x 1032 can
make use of this offset and gain correction images for real-time
shading correction purpose.

How to switch shading correction on


1) Select the Image > Configure Input... > Input tab.
2) Select the Shading correction > Offset check box to have the camera’s
offset image automatically subtracted during image acquisition.
3) Select the Gain check box to have each image acquired automatically
divided by the camera’s gain image.
Activating shading
correction.
36

4) Click on OK.
" Now all images you are acquiring via the current input channel will
be shading corrected.
Defining gain and offset correction

How to check reference images


1) Set electron beam intensity at medium brightness.
" Generally, after having acquired reference images correctly, you
will not need to adjust current brightness. During the subsequent
image acquisition, the online histogram ought to display a mean
value between 40 % and 60 %.
2) Acquire an image - without specimen - using the Image > Acquire or
Snapshot command.
" The corrected acquisition cannot have any visible structures due to
the imaging system. The only thing you should be able to see is
homogeneous noise.
A microscope image
acquired without specimen
shows the homogeneous
noise visible when the
shading correction was
conducted successfully.
This noise is caused by the
electrons’ quantum noise.
The image (right) has been
contrast enhanced.

" If shading correction was not conducted successfully, please check


and see whether the reference images acquired look essentially the
same as the sample images shown here. To do this, use both
Image > Acquire reference images... > Show buttons. Then - if
necessary - repeat acquiring reference images for shading
correction. Be sure to follow all instructions as described above.
3) Check the exposure time in the Camera Control dialog box to be the

37
standard value of the current input channel.
" The Camera Control dialog box features the control lamp Ref.. It is
white when there are no reference images defined which have the
same image format as the current input channel.
" The control lamp is green when the exposure time is within a time
range of about 20% of the exposure time you have used to acquire
the reference images with. This guarantees, that the reference
images are valid to correct the shading artifacts.
" The control lamp is red when the exposure time is to far out of the
value you have aquired the reference images with. In this case the
shading correction will be applied but it can’t be guaranteed that the
artifacts will be removed completely. Adjust the exposure time until
the lamp is green.
MegaViewIII • Camera set-up under analySIS

" If you need a different exposure time for standard, you have to
acquire new reference images which refer to this exposure time.
4) Make sure that the focus is correct before acquiring reference images.
You should now repeat the whole process for other image formats required,
e.g. 688 x 516 with 2x2 binning which is typically used for focus or search
mode. But keep in mind, that online shading correction reduces the speed
of the image acquisition.

Calibrating input channel


Calibration should be done for every input channel that is used to make
image snapshots.

How to calibrate an input channel


1) Insert a calibration standard sample into your microscope. This is
normally a grid or grating with regular known intervals (e.g. a replica
with 2160 lines/mm or 463 nm line spacing).
2) Select the Image > Set Input command. Select the input channel you
want to calibrate, usually a snapshot channel.
3) Set a reasonable microscope magnification.
4) Use the Image > Acquire command to optimize the acquisition
parameters of the calibration sample.
5) Select the Image > Configure Input... command and activate the XY
Calibration tab.
38

related topics
Focusing the camera p. 44
Calibrating input channel

Calibrating input channel.

6) Enter this magnification level into the Magnification field.


7) Enter the length you want to calibrate with into the Calibration length
field.
8) Click on the Unit... button.
" Select m (for meters) in the Basic unit list in the Set Unit dialog box.
" Select, eg, n in the Scale list if you wish the calibration length to be
nm.
9) Select the Arbitrary option in the Calibration group.
10) Leave the X/Y ratio for the MegaViewIII camera at 1.
11) Click on the Calibrate button.
" The mouse cursor will appear within the active image.
" Position the mouse cursor at the starting point of the calibration

39
length and leftclick.
" Then position the mouse cursor at the final point of the calibration
length and confirm by leftclicking. The line must be the exact same
length as the calibration length you entered.
12) Click on the Save... button to open a list of calibration data for various
magnifications.
13) Click on the Add button to add the data of the calibration you’ve just
completed to the magnification list and have it included in the diagram
as well.
14) If the list in the Magnification Table dialog box initially included invalid
calibration data then select these entries and click on the Delete button.
MegaViewIII • Camera set-up under analySIS

Defining calibration curve.

15) Exit this dialog box by clicking on OK.


16) Repeat this for other magnifications to obtain a magnification
calibration curve.
" To do so, you usually don’t have to leave the Configure Input dialog
box. Alter the microscope magnification. Click on the Acquire button
in the Configure Input dialog box and use the live image onscreen
to position and focus the calibration specimen. When you’re
satisfied, quit the live mode by clicking on Snapshot. Now you can
repeat the calibration on that image.
17) When you have finished calibration, confirm it via OK.
Note When you have calibrated an existing channel, you don’t have to calibrate
new channels. You can transfer the calibration data of the existing channel
to any new channel.
To do so, select the Image > Set Input... command. Activate the existing
40

channel whose calibration data you want to transfer. Press the New channel
button. In the Select device dialog box, choose the MegaView III FW entry.
Select the Use calibration of active channel check box and confirm via OK.

Activating the remote control


Some microscopes have remote control capabilities that would allow
analySIS® to read the current magnification of the microscope via a serial
cable connection. This ensures that all images acquired will carry the
correct calibration information assuming that the microscope or input
channel is properly calibrated.

How to activate the remote control


1) Select the Image > Configure Input... command and activate the
Magnification tab.
Magnification setting

Activating remote.

2) The name of the microscope should be displayed in the Device list.


Select this device.
3) Select the Remote > On check box.
4) Try to get a magnification reading by clicking on the Read button.
" Magnification is automatically read via the remote every time a
snapshot is made.

Magnification setting
If you do not have remote features on your microscope, then you are
required to set the magnification of the image whenever a snapshot is made.
1) Select the Image > Configure Input... command and activate the Macro

41
tab.
2) Type in the function SetMagnification$() in the Postprocessing steps
field.
" This makes sure that you will not forget to set the magnification of
the images that you acquire. The Set Magnification dialog box will
pop-up after each image snapshot.
MegaViewIII • Camera set-up under analySIS

3) Confirm your input calibration by clicking on OK.


4) Click on the Standard (button bar) > Snapshot button.
" The live image will be frozen.
" The Set Magnigication dialog box will be opened. Enter the
microscope magnification and confirm via OK.
42
Servicing Notes

Servicing Notes
Warning The instructions in this sections are for service technicians only. If you
encounter any problems while installing the camera, please contact our
support desk.
Warnings! • Always remember to move the MegaViewIII to the OUT position (out of
beam path) before removing the pressurized air supply from the
MegaViewIII controller box. The green button of the In/Out position
control box should be lit. If the MegaViewIII was left in the IN position,
you may have an air leakage when you reconnect the air supply.
• The air throttle valves on the MegaViewIII unit are pre-adjusted for
normal in/out movement speed. Opening the valves too much may
cause the unit to move too fast thus causing mechanical damage. If
adjustments are required, do it in small increments and always start
from the closed position.

Adjusting the MegaViewIII In/Out movement speed


How to adjust the MegaViewIII In/Out movement speed
Warning You are normally not required to disassemble any part of the MegaViewIII
system upon installation except for the purpose of focusing the lens system.
Therefore these notes are only necessary if there is a need to service the
unit.
1) The air throttle valve that controls the IN position movement is the one
nearer to the flange, the valve that controls the OUT position
movement is the closer to the camera.
2) Closing a valve will cause movement to slow down and opening a valve
will increase the speed. If the valve is closed too much, it may not
move at all. If the valve is opened too much, it may move too fast. You
should adjust the valves such that it takes approximately 2 seconds to
move between positions.

43
Adjusting air throttle valves
that control the IN or OUT
movement speed.

3) To be able to open or close the valve, first loosen the security nut by
turning it counterclockwise. This releases the valve control screw so
that it may be turned.
MegaViewIII • Servicing Notes

First loosen the security nut


to be able to turn the valve
control screw.

4) Turn the valve control screw clockwise to close the valve – this slows
the movement speed.
Turn the valve control screw anti-clockwise to open the valve – this
increases the movement speed.
5) Tighten the security nut by turning it clockwise. The security nut
prevents the valve from accidental opening or closing.

Adjusting the prism position


Warning You are normally not required to disassemble any part of the MegaViewIII
system upon installation except for the purpose of focusing the lens system.
Therefore these notes are only necessary if there is a need to service the
unit.
Note The prism position varies with the model of microscope. The camera is
delivered having the prism position preadjusted to your type of microscope.
So usually, you don’t have to change the prism position.
when is this necessary? You only have to adjust the prism position when the shadow of the prism on
the microscope fluorescence screen is not symmetric.
This is how the shadow of
the prim should look like on
the fluorescence screen of
the microscope.
44

How to adjust the prism position


1) The prism can be moved by several millimetres. This adjustment is
typically done to centre the prism in the field of view of the microscope.
2) At the bottom of the prism, there are 2 hex lugs holding the prism to the
two rods, called prism holding lugs. Loosen these lugs and slide the
prism to the desired position, then tighten again. Keep the face of the
prism parallel to the face of the flange.
3) Follow the procedure in the next section to refocus the lens of the
camera.
Focusing the camera

Focusing the camera


Warning You are normally not required to disassemble any part of the MegaViewIII
system upon installation except for the purpose of focusing the lens system.
Therefore these notes are only necessary if there is a need to service the
unit.

How to focus the camera


1) Focusing of the camera unit should be done when the system is
running. On some models of microscope a beam stop is available
above the scintillator plane and can be easily used for focusing. If not,
a grid with a convenient specimen should be inside the TEM to focus
on.
2) Remove the elliptical housing. There are two small screws located at
the bottom of the housing.
Remove the two housing
screws and remove the
camera housing.

3) Check the distance from front of the flange surface to the middle of the
prism. This distance varies with the model of the microscope.
4) Check that the prism holding lugs and the prism rod holding lugs

45
are sufficiently tight.
5) Use an input channel with the following settings:
" 688x516 image size format with 2x2 binning,
" Exposure time - 50 ms,
" Histogram display - off,
" Automatic gain display - on,
" Real time FFT - on,
" Shading correction - on.
" The respective offset and gain images ought to be acquired.
Warning! Do not loosen the prism rod holding lugs because the vacuum of the
microscope could suck the prism with the rods into the column, with
disastrous consequences.

related topics
Defining gain and offset correction p. 29
MegaViewIII • Servicing Notes

Prism rod holding lugs. DO


NOT LOOSEN!

6) Loosen the camera holding lugs.


Camera holding lugs.

7) Get a best focus image of your sample on the microscope fluorescent


viewing screen.
8) Move the MegaViewIII unit into the beam path.
9) Loose the fixing ring.
Loose the fixing ring. Now
you can turn an Allen key
within a focusing hole and
thus change the focus
length.
46

10) Put an regular L shaped Allen key into one of the focusing holes and
turn the front of the lens with the Allen-key to change the focus length.
" You can focus more precisely using the RT-FFT spectrum. A best
focus image has the maximum spectral spread (more high
frequency information), i.e., the higher the distance of an image
structure to the center the better. Keep the front of the lens against
the frame always. Remember to always use the largest lens
aperture.
Focusing the camera with beam stop

The fourier image of a 2-D


catalase crystal.
The light spots represent the
periodic structures in the
microscope image. The
distance of a spot to the
center increases, when the
camera is better focussed.

11) When satisfied with the focus, tighten the fixing ring and the camera
holding lugs.

Focusing the camera with beam stop


When your microscope has a beam stop, you can more easily make use of
it instead of a specimen.
Beam stop

47

How to focus the camera using beam stop


1) Use the MegaView III Focus channel.
" The respective offset and gain images for shading correction must
be available.
2) Start the live modus.
3) Move in the beam stop.
" Use the RTFFT spectrum to focus the camera. The camera is best
focussed, when the indictated Fourier structure reaches the edge
of the RT-FFT window.
MegaViewIII • Servicing Notes

The structure indicated by


the arrow ought to reach the
edge of the RT-FFT window.
48
Appendix

Appendix
Technical specification
Image resolution Standard: 1376 x 1032 pixels
2x binning: 688 x 516 pixels
Binning 2x, 4x, 8x
Pixel size 6.45 x 6.45 µm, virtual about 20 x 20 µm
Pixel clock rate 20MHz @ 12 bit
Dynamic range 4096 gray values, 12 bit
Exposure time 100 µs - 160 s
Frame rate (depending on reso- > 10 fps at standard resolution
lution and PC speed) > 20 fps at 2x binning
Display Full image at any frame rate
Cooling element 10°C, Peltier cooled
Temperature monitor CCD chip & Housing
Readout noise Correlated Double Sampling
Camera mount One flange at wide angle port
Camera positioning PC controlled pneumatic movement / Control
box
Camera coupling Lens-optically coupled
Anti-Blooming ≥ 300
Interface FireWire TM (IEEE1394)

Packing List
1) MegaViewIII camera unit assembly.
2) Flange screws.
3) MegaViewIII controller box.
4) MegaViewIII In/Out position control box (handbox).
5) analySIS ® PC with software and FireWire TM board installed. (If
complete system is delivered.)
6) FireWireTM cable (PC to camera).

49
7) DB9-DB9 serial cable (PC to MegaViewIII controller box).
8) DB9-DB9 serial cable (MegaViewIII In/Out position control box to
MegaViewIII controller box)
9) 2x bare pneumatic tubes (5mm ∅). (MegaViewIII controller box to
MegaViewIII camera unit).
10) 1x pneumatic tube (6mm ∅) with connector to self-closing valve on one
end.
11) 1x pneumatic tube (6mm ∅) with self-closing valve on one end and T-
joint on the other end. The size and type of T-joint may vary for the type
of microscope of pneumatic system in use.
12) analySIS ® software package (or included in item 5).
13) User’s guide
14) This installation guide.
Index

A O
Acquire Images 30 Offset image 32
Adjust Online shading correction 31
In/Out movement speed 43
Prism position 44
P
Prism holding lugs 6
B Prism rod holding lugs 6
Beam stop 47

Q
C Quick-start installation 2
Cables 10
Camera cable
Connect to computer 14 R
Camera Control 30
Check reference images 37 Remote control 40

F S
FireWire board IEEE-1394 4 Servicing Notes 43
Installation 5 Set Magnification 41
Focus camera 45 Snapshot 30
Software Installation
Windows 2000 12
Windows XP 21
G
Gain image 33
W
Windows 2000
I Software Installation 12
Input Channel 29 Troubleshooting 15
Calibrate 38 Windows XP
Software Installation 21

M
MegaViewIII Controller 4
MegaViewIII controller unit
Installation 8
MegaViewIII unit 6

91
CE DECLARATION
OF CONFORMITY

We Soft Imaging System GmbH


Hammer Str. 89
D-48153 Münster
Name and Address of manufacturer or representative

declare, according to legal obligation, that the following product complies with the protection
requirements of the 89/336/EEC council directive in accordance with the laws of member states
relating on electromagnetic compatibility, as amended by directive 93/68/EEC:

digital CCD camera MegaView III *


Type

The product to which this declaration refers is in conformity with the following relevant and
harmonized standards, the reference numbers of which have been published in the Official Journal
of the European Community:

EN 50082-1:1997
EN 55022:1998+A1:2000, cl. B
EN 55024:1998
EN 61000-4-2:1995+A1:1998
EN 61000-4-3:1996+A1:1998
ENV 50204:1995
EN 61000-4-4:1995
EN 61000-4-6:1996

*tested using SIS PC

Date: November 30th, 2002

Dr Manfred Schierjott April 30th, 2002


Date