Olimpus FLUOVIEW FV500 User manual

User’s Manual
FLUOVIEW
FV500
CONFOCAL LASER SCANNING
BIOLOGICAL MICROSCOPE
Ver.3.3
This user’s manual is for the software to be run on Olympus FLUOVIEW FV500 Confocal Laser
Scanning Biological Microscope. To ensure safety, obtain optimum performance and familiarize yourself
fully with this product, we recommend that you study this manual thoroughly before operation.
This user’s manual is composed of two volumes including “SYSTEM OVERVIEW”, “OPERATION
INSTRUCTIONS”, “MAINTENANCE” and “TROUBLESHOOTING”. Together with this manual, please
also read the “User’s manual FLUOVIEW FV500” and the instruction manual of the microscope in order
to understand overall operation methods. To ensure the safety operation of laser system, we
recommend you to study the manual of each laser and the light source equipment besides this manual.
Retain this manual in an easily accessible place near a system for future reference. AX9787
Petition

1
Page
CAUTION
1. Reproduction, copying or duplication of a part or all of this software and manual is prohibited.
2. The information described in this manual may be subject to change without notice.
Registered Trademarks
Microsoft, Microsoft Windows, Excel for Windows are registered trademarks of Microsoft Corporation.
Other brand names and product names are trademarks or registered trademarks of their respective owners.

FLUOVIEW MANUAL CONFIGURATION
2Page
FLUOVIEW MANUAL CONFIGURATION
The FLUOVIEW system uses two manuals including this “User’s Manual” and the on-
screen manual built into the software (“Online Help”).
The User’s Manual is composed of the five following volumes and subject matter:
•SAFETY GUIDE (Anoter manual)
Describes notes and cautions on using the FLUOVIEW system and on types of
warning labels.
•SPECIFICATIONS (Another manual)
Describes the specifications of the FLUOVIEW system.
•SYSTEM OVERVIEW (This manual)
Describes the outline of the FLUOVIEW system.
•OPERATION INSTRUCTIONS (This manual)
Describes the operation procedures of the FLUOVIEW system, for example,
methods for image acquisition and various image processing.
•MAINTENANCE (This manual)
Describes maintenance of the FLUOVIEW system.
•TROUBLESHOOTING (This manual)
Describes countermeasures in case trouble occurs.
For Online Help, please see “1-3 Online Help” in “OPERATION INSTRUCTIONS” of this
manual.

NOTATIONS IN THIS MANUAL
3
Page
NOTATIONS IN THIS MANUAL
This manual complies with the following notations.
Notation of Caution, Notes and Tips
Notation Description
Caution to prevent injuries to the user or damage to the product
(including surrounding objects).
NOTE
Note for the user.
TIP
Hint or one-point advice for user reference.
Notation of panel, Command Buttons and Dialog Boxes
Notation Description
[Acquire] panel The name of a panel, dialog box, list box or check box is
enclosed inside square brackets ([ ]).
<OK> button
<Open File> button
The name of a button in a panel or dialog box is enclosed
inside angled brackets (< >).
Notation of Mouse Operations
Notation Description
clicking Action of pressing, then immediately releasing the mouse
button.
double-clicking Action of clicking the mouse button twice in quick succession.
dragging Action of moving the mouse while holding down the mouse
button, then releasing the mouse button at the desired
destination.
(Note) In this manual, clicking, double-clicking and dragging involves pressing the left
button of the mouse, unless otherwise specified.

NOTATIONS IN THIS MANUAL
4Page
Notation of key operations
Notation Description
Enter The name of a key is enclosed inside .
F1
+
Alt The positive sign (+) expresses the combination of more than
one key operation.
For example, F1
+
Alt refers to pressing the
F1 key while holding the Alt key down.
Direction keys Generic names given to the →, ←, ↑and
↓keys.
Notation of system-specific terms
Notation Description
XY observation
(Other observations)
Refers to observation with XY scanning.
(The same principle also applies to other observations such
as XZ, Xt, XYZ, XYt and XYZt.)
Note that some of the panels and dialog boxes shown in this manual are not the precise
reproductions of the originals. Some windows are resized to facilitate the reading and
some grayed-out characters are printed in readable characters.

This volume describes the outline of the FLUOVIEW FV500
system.
Please read this volume so that you can understand the
system before use.
On This Volume
S
SY YS ST TE EM M O OV VE ER RV VI IE EW W

CONTENTS
1 SYSTEM OVERVIEW 1-1
1-1 Principle ............................................................................................1-1
1-2 Features of FLUOVIEW FV500 ........................................................1-2
1-3 Light Path Diagram ..........................................................................1-3
1-4 System Configuration ......................................................................1-4
1-4-1 System Component Units and Their Functions ........................................... 1-4
1-4-2 Block Diagram............................................................................................. 1-6
1-5 Software Functional Configuration ................................................1-9
1-5-1 Function Panel and Display Panel............................................................... 1-9
1-5-2 Panel Structure of the Software................................................................. 1-10
1-5-3 Icons Executed by Dragging & Dropping................................................... 1-11
1-5-4 Identification of Images Depending on the Observation Methods.............. 1-12

SYSTEM OVERVIEW/Principle
1-1
Page
1 SYSTEM OVERVIEW
Olympus FLUOVIEW FV500 is a confocal scanning type laser reflected fluorescence microscope which uses
confocal optics for high resolution, high contrast and drastically improved resolution in the light axis direction.
It offers researchers features for sectioning, 3D construction and time-series observation as well as a variety of
image processing and analysis functions.
This section describes the principle and characteristics of the system and the functional configuration of
software for use with the system.
1-1 Principle
A laser scanning microscope converges a laser beam into a very small spot using an
objective and scans the specimen in the X-Y directions.
It then detects the fluorescence and transmitted light from the specimen using light
detector and outputs the image of the
specimen on an image monitor.
The confocal optics place a confocal
aperture at a position which is optically
conjugate with the focusing position (i.e.
confocal plane) to eliminate light from
parts other than those for the focusing
position. As a result, the part
corresponding to the eliminated light is
darkened in the image, making it
possible to slice a thick tissue
specimen optically.
On the other hand, with ordinary optical
microscopes, light from a part other
than the focusing position overlaps with
the image-forming light on the focusing
position, so the overall image tends to
be vague.
Transmitted light images can be
obtained by use of a transmitted light
detector, but the images will not be
confocal images, because there is no
confocal aperture in front of the detector.
The transmitted light image can be used by combination with a confocal fluorescence
image to observe fluorescent localization.
Confocal aperture
Light detector
Objective
Specimen
Light detector
Laser

SYSTEM OVERVIEW/Features of FLUOVIEW FV500
1-2 Page
1-2 Features of FLUOVIEW FV500
1. The detector has 12-bit resolution and is capable of detecting very small changes in
fluorescence inside cells.
2. The system has high resolution of 2048 x 2048 pixels. The output video signal is
non-interlaced for clear images without flickering.
3. The transmitted light is detected by a photomultiplier for bright, sharp transmitted
images.
4. The auto gain adjustment feature eliminates the need for complicated gain
adjustment operation.
5. A total of 5 image channels, including up to 4 fluorescence images and a transmitted
image, can be acquired simultaneously.
6. The scan unit can be used in common with an upright or inverted microscope.
7. The confocal aperture is continuously variable and the optimum confocal aperture for
each objective is set automatically.
8. High-speed scanning acquires 4 images per second (512 x 512 pixels).
9. Sequential scan for acquiring images without crossover.
10.In addition to raster scanning, scanning modes such as vector scanning and oblique
scanning are provided to meet a wide range of applications.
11.The dichroic mirror for excitation, dichroic mirror for spectroscopy, confocal aperture
and emission filters are set automatically.
12.When using the system with AOTF (FV5-COMBA), the following functions are
available.
•Line Kalman scanning
•Line sequential scanning
•Image acquisition in the REX/Bleach mode(FRAP experiment)

SYSTEM OVERVIEW/Light Path Diagram
1-3
Page
1-3 Light Path Diagram
Emission DM turrets 1 to 3
Pinholes 1 to 4
Confocal
lenses 1 to 4
Emission filter turret
Mirror
To IR laser unit
To UV laser unit
To VIS laser
combiner
Mirrors
DM
X/Y galvano
Mirrors
Excitation
DM turret

SYSTEM OVERVIEW/System Configuration
1-4 Page
1-4 System Configuration
1-4-1 System Component Units and Their Functions
Photo 1-1 Combination with AX70
Scan unit
Heart of a laser microscope, composed
of the light scanner and light detector.
Microscope
AX70 upright microscope set up
for fluorescence observation.
Multi-control box
Controller of the AX70 microscope.
Computer
Used to control the
LSM, process image
files, etc.
Transmitted light detector
unit
Unit for acquiring transmitted
images. Connected with the
microscope through an optical
fiber cable.
Laser combiner
Unit for merging laser beams from the lasers
such as an Ar laser, Kr laser and HeNe laser
into a single beam to be transmitted through
the optical fiber cable.
Control unit
Unit for driving the
scanning unit, laser
combiner, X-motor
and transmitted light
detector.
Image monitor
Video monitor for
displaying laser
scanning images,
control panel, etc.
Transmitted
light unit
Unit
incorporating a
100 W halogen
lamp bulb to
provide
illumination for
transmitted light
observation.
Connected with
the microscope
through an
optical fiber
cable.
Reflected light unit
Unit incorporating a
100 W mercury
burner to provide
illumination for
reflected light
observation.
Connected with the
microscope through
an optical fiber cable.
Power box
Power supply unit
for the AX70
microscope.
Anti-vibration
platform
Exclusively
designed
pneumatic
platform.
Reflected Fluorescence
Light Power Supply Unit
Power supply unit for the
mercury burner for reflected
light observation.
Z-motor
Motor for driving the
focusing block of the
microscope
Laser power
supply units

SYSTEM OVERVIEW/System Configuration
1-5
Page
Photo 1-2 Combination with IX
Microscope
Upright microscope exclusively
designed for FLUOVIEW FV500.

SYSTEM OVERVIEW/System Configuration
1-6 Page
1-4-2 Block Diagram
50pin
(5)
2.9m
50pin
Dsub15pin
(8)
2.9m
(1)
2.9m
Dsub15pin
(1)
2.9m
(2)
2.9m
Ar LASER POWER
SUPPLY UNIT
Kr LASER POWER
SUPPLY UNIT
HeNe-G LASER
POWER SUPPLY
UNIT
HeNe-R LASER
POWER SUPPLY
UNIT
1.8m
1.8m
5.5m
5.5m
FV5-COMB
LASER COMBINER
(10)
2m
MICRO-
SCOPE
Dsub15pin
BNC
(3)
3m
Dsub15pin
(2)
2.9m
FV5-SU
SCAN UNIT
Dsub25pin
Dsub25pin
(1)
2.9m
(1)
2.9m
Dsub9pin
(3)
2m
FV5-ZM
Z-MOTOR
FV5-TD
TRANSMIT
TED LIGHT
DETECTOR
LG-PS2
TRANSMITTE
D LIGHT UNIT
Dsub25pin
FV5-PSU
POWER
SUPPLY
UNUT
(7)
2.9m
(4)
2.9m
BNC
(1)
2.9m
MONITOR
COMPUTER
(4) 2.9m
U-MCB
(combination
with AX only)
Dsub9pin
Dsub9pin
Dsub9pin
Dsub9pin
IR LASER
POWER SUPPLY 2m
3m
Mercury
burner
U-RFL-T /
BH2-RFL-T3
Reflected
Fluorescence
Light Power
Supply Unit
CHILLER
FV5-LUUV
FV5-LDUIR
REMOTE SW
(3)
1m
(10)
(2)
2.9m
(10)
1.7m
(1)
2.9m
(1)
(1)
(2)
(6)
2.9m
Square, 3-pin
HeCd LASER
POWER SUPPLY
1.8m
Kr LASER
SIROCCO FAN
Duct, 2.2m
UV-Ar LASER
POWER
SUPPLY
(2)
2.9m
(10)
2m
(7)
2.9m
U-PS
(combination
with AX only)
FV5-LUHECD
(3)
3m
(2)
2.9m
(1)
2.9m
Circular,8-pin
Dsub25pin
Dsub15pin
Dsub15pin
Dsub25pin
(1)
2.9m
(11)
(11)
Dsub9pin
Square, 6-pin
Circular,6-pin
Circular,37-pin
Dsub25pin
Circular,37-pin
Square, 2-pin
Square, 2-pin
(1)
2.9m
Dsub25pin
Square, 2-pin
(10)
2.2m
(10)
2.2m
Dsub15pin
BNC
Dsub25pin
Circular,7-pin
Square, 3-pin
Dsub25pin
Square, 3-pin
Dsub25pin
Circular,37-pin
Square, 3-pin and
Circular 8-pin
Circular,37-pin
(10)
Dsub25pin
Dsub9pin
BX-UCB
(combination with
BX51,61 only)

SYSTEM OVERVIEW/System Configuration
1-7
Page
The block diagram in AOTF (FV-COMBA) use.
50pin
(5)
2.9m
50pin
Dsub15pin
(8)
2.9m
(1)
2.9m
Dsub15pin
(1)
2.9m
(2)
2.9m Ar LASER POWER
SUPPLY UNIT
HeNe-G LASER
POWER SUPPLY
UNIT
HeNe-R LASER
POWER SUPPLY
UNIT
1.8m
1.8m
5.5m
5.5m
FV5-COMBA
LASER COMBINER with AOTF
(10)
2m
MICRO-
SCOPE
Dsub15pin
BNC
(3)
3m
Dsub15pin
(2)
2.9m
FV5-SU
SCAN UNIT
Dsub25pin
Dsub25pin
(1)
2.9m
Dsub9pin
(3)
2m
FV5-ZM
Z-MOTOR
FV5-TD
TRANSMIT
TED LIGHT
DETECTOR
LG-PS2
TRANSMITTE
D LIGHT UNIT
Dsub25pin
FV5-PSU
POWER
SUPPLY
UNUT
(7)
2.9m
(4)
2.9m
BNC
(1)
2.9m
MONITOR
COMPUTER
(4) 2.9m
U-MCB
(combination
with AX only)
Dsub9pin
Dsub9pin
Dsub9pin
Dsub9pin
IR LASER
POWER SUPPLY 2m
3m
Mercury
burner
U-RFL-T /
BH2-RFL-T3
Reflected
Fluorescence
Light Power
Supply Unit
CHILLER
FV5-LUUV
FV5-LDUIR
REMOTE SW
(3)
1m
(10)
(2)
2.9m
(10)
1.7m
(1)
2.9m
(1)
(1)
(2)
(6)
2.9m
Square,
3-pin
HeCd LASER
POWER SUPPLY
1.8m
Kr LASER
SIROCCO FAN
Duct, 2.2m
UV-Ar LASER
POWER
SUPPLY
(2)
2.9m
(10)
2m
(7)
2.9m
U-PS
(combination
with AX only)
FV5-LUHECD
(3)
3m
(2)
2.9m
(1)
2.9m
Circular,8-pin
Dsub25pin
Dsub15pin
Dsub15pin
Dsub25pin
(1)
2.9m
(11)
(11)
Dsub9pin
Square, 6-pin
Circular,6-pin
Circular,37-pin
Dsub25pin
Circular,
37-pin
Square, 2-pin
Square, 2-pin
(1)
2.9m
Dsub25pin
Square, 2-pin
(10)
2.2m
(10)
2.2m
Dsub15pin
BNC
Dsub25pin
Circular,7-pin
Square, 3-pin
Dsub25pin
3-pin
Dsub25pin
Circular,37-pin
Square, 3-pin and
Circular 8-pin
Circular,37-pin
(10)
Dsub25pin
Dsub9pin
FV5-LCU
Laser Control
Unit
1m
Kr LASER POWER
SUPPLY UNIT
1m 50pin
Circular
X2
(1)
2.9m
BX-UCB
(combination with
BX51,61 only)

SYSTEM OVERVIEW/System Configuration
1-8 Page
Either the Kr laser or HeNe-G laser can be used.
(1) Control cable
(2) Interlock cable
(3) Optical fiber cable
(4) Analog signal cable
(5) I/O cable
(6) Galvano control cable
(7) Galvano X/Y cable
(8) Power/control cable
(9) Power cable
(10) Laser power cable
(11) RS-232C cable
Input and output terminals on FV5-LCU back
Outputs for laser intensity
(Analog, 0-10V)
(VIS1 and VIS2 are for
invisible laser, UV is for UV
laser, IR is for IR laser and
Blanking is for switching laser
ON/OFF.)
(UV laser and IR laser are
optional.)
For connection to PC (PC-
SYSTEM), 50 pin.
For connection to FV5-
PSU (PC-SYSTEM), 50
pin.
For switching signals
output from the left
connectors
(VIS1 and VIS2 are for
invisible laser, UV is for UV
laser, IR is for IR laser and
Blanking is for switching
laser ON/OFF.)
(UV laser and IR laser are
optional.)
VIS1 VIS2 UV IR
VIS1VIS2 UV IR Blanking Event output signal 1 (TLL)
Event output signal 0 (TLL)
Trigger input signal 1 (TTL)
Trigger input signal 0 (TTL)
Horizontal sync signal (TTL)
Vertical sync signal (TTL)
For connection to FV5-TD
(Dsub 25 pin, male terminal)
For connection to FV5-PSU
(TD)
(Dsub 25 pin male terminal)
For connection to (AOTF) FV5-
COMBA
(Coaxial connector)
Service connector

SYSTEM OVERVIEW/Software Functional Configuration
1-9
Page
1-5 Software Functional Configuration
This software uses panel-type windows.
Usually, it is required to “select a menu then select the command to be executed” in
order to execute a function provided by software. With the panel system, a software
function can be executed easily by “selecting the panel page tab of the function to be
executed”, just like when using a system notebook or file folder.
1-5-1 Function Panel and Display Panel
The FLUOVIEW software is organized by two kinds of panels, the function panels and
display panel.
The function panels include the [Acquire], [File I/O], [Tile], [Process], [Analyze] and
[Visualize] panels.
The display panel shows either the [Live] panel or the panel image loaded from a file
([(filename)] panel).
In this manual, the function panels are referred to simply using their
page tabs.
Namely, the [File I/O] panel of function panel is referred to simply as
the [File I/O] panel.
NOTE
Function panel Display panel

SYSTEM OVERVIEW/Software Functional Configuration
1-10 Page
1-5-2 Panel Structure of the Software
This software cannot show the indexes of all function panels at a glance, but uses
scrolling to display the desired page tab. Please use the following list of the panels for
reference in scrolling.
Acquire
Scan
Z Stage
Sets up the image acquisition and executes actual acquisition.
Sets the zooming ratio and observation mode for image acquisition.
Sets the Z-direction scanning range for image acquisition.
Time Series Sets the interval period for image acquisition.
Optics Sets the microscope light path for image acquisition.
File I/O
Tile
Process
Math
Filters
Histogram
Logical
Analyze
Single
Saves, loads and deletes images.
Changes the image display method.
Processes the acquired images.
Performs mathematical operations between images.
Filters images.
Changes the image contrast.
Performs logical operations between images.
Analyzes image data.
Obtains the intensity values, intensity distribution, length, area and
average intensity in images.
Orientation
Other Options
Series Obtains the change in the sum of intensity values in images.
Visualize Constructs an image from a different viewpoint or displays a 3D image.
Sets the image rotation angle, direction and number.
Sets the various 3D Rendering.
Dyes Sets the fluorescence dye method for image acquisition and reagent for
each channel.
Lasers Sets the ND of laser for image acquisition.