iCycler iQ
™
Multi-Color
Real Time PCR Detection
System
Upgrade Installation Guide
Catalog Number
170-8744
For Technical Service Call Your Local Bio-Rad Office or in the U.S. Call 1-800-4BIORAD (1-800-424-6723)
Table of Contents
Page
Section 1 System Component/Packing List ...............................................................1
Section 2 Installing Filters ...........................................................................................2
Section 3 Loading Software and Firmware...............................................................4
Section 4 Adjusting the Mask in Imaging Services...................................................6
4.1 Imaging Services ........................................................................................................6
4.1.1 Description ..........................................................................................................6
4.1.2 Adjusting the Masks............................................................................................7
4.1.3 Checking Mask Alignment .................................................................................8
4.1.4 Image File............................................................................................................8
Section 5 Quick Guide to Single or Multi-Color Experimentation ........................9
5.1 Well Factors .............................................................................................................10
5.1.1 Well Factor Source: Experiment Plate .............................................................10
5.1.2 Using the Experimental Plate for Well Factors................................................11
5.1.3 Well Factor Source: Well Factor Plate .............................................................12
Section 6 Quick Guide to Protocol Workshop.........................................................13
6.1 Creating a New Protocol ..........................................................................................13
6.2 Editing a Stored Protocol .........................................................................................14
6.3 Creating a New Plate Setup .....................................................................................16
6.4 Editing a Stored Plate Setup.....................................................................................17
Section 7 Quick Guide to Collecting and Analyzing Data .....................................18
Section 1
System Components/Packing List
The Multi-Color Upgrade kit for the iQ Optical Module is shipped with the following
components:
• CD-ROM with iCycler iQ Software
• Upgrade Installation Guide
• Multi-Color Optical System Instruction Manual
• Texas Red™/Rox Filterset
Includes: 575/30X Excitation Filter and 620/30M Emission Filter
• iCycler iQ Dye Calibration Solutions
Includes: Calibrator solutions for FAM, TET, HEX, JOE, TAMRA, Texas Red,
Cy™3 and Cy™5 and External Well Factor Solution
Texas Red is a registered trademark of Molecular Probes, Inc.
Cy is a trademark of Amersham Pharmacia Biotech.
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Section 2
Installing Filters
The iQ detector is shipped with a set of filters for fluorescein are in position 2 of both
filter wheels, and the remaining positions are filled with filter blanks. All filters are mounted
in filter holders (see Figure 2.1). Each position on both filter wheels must have either a filter
or filter blank installed in order to avoid possible damage to the detector. Position 1 of both
filter wheels is dedicated to filter blanks and may not be changed.
You should insure that the filters and blanks are in their correct positions.
FilterSet4
Position
Excitation Emission Recommended Fluorophores
2 490/20X 530/30M Fluorescein (FAM),SYBR Green
3 530/30X 575/20M HEX, TET, VIC, JOE, Cy3
4 548/25X 595/24M TAMRA
5 575/30X 620/30M Texas Red, ROX
6 635/30X 680/30M Cy5, LC640
FilterSet5
Position Excitation Emission Recommended Fluorophores
2 490/20X 530/30M Fluorescein (FAM),SYBR Green
3 530/30X 575/20M HEX, TET, VIC, JOE, Cy3
4 53030X 620/30M Ethidium Bromide
5 610/30X 660/20M LC640
6 660/30X 710/20M Cy5.5, LC705
Note: The filter designation 490/20X indicates that this filter will allow light from 480/500 nm
to pass through. The first number, 490, indicates the center of the wavelength of light. The
second number, 20 indicates the total breadth of wavelengths of light that can pass through it.
X indicates excitation only and M indicates emission only types of filters. Excitation and
emission filters are not interchangeable.
Fig. 2.1. Filter in filter holder.
TOP VIEW
SIDE VIEW
Ta b
Ta b
Filter or
Filter Blank
1. Turn off the power to the Optical Module.
2. Release the two black latches on each side of the Optical Module. Slide the housing
backwards 2–3" (5–8 cm), exposing a black case, the filter wheel housing. It is not
necessary to remove the housing or cables.
3. Remove the two rubber plugs on the top of the filter wheel housing by pulling them
straight upward. These plugs shield the filter wheels. The excitation filters are located in
the slot on the right side of the instrument; the emission filters are located in the slot in the
center of the instrument (see Figure 2.2). The tops of the filter blanks in position 1 (the
"home" position) should be visible in the slots containing both the excitation and
emission filters; the number "1" should be visible in the slot on each side of both filters.
Changing both types of filters is similar.
4. Turn the filter wheels to the desired positions using the ball end hex driver. As long as the
power to the Optical Module is off, the filter wheels may be turned freely in either direction.
5. To remove a filter, grasp it on both sides with the filter removal pliers and squeeze the tab
in; gently pull the filter up and out.
6. To insert a filter, grasp the filter with the pliers and insert it into a vacant slot. For the
excitation filters, the tab on the filter is toward the front of the instrument. For the
emission filters, the tab on the filter is toward the right of the instrument. Be sure that
every position in the filter wheel has either an excitation or emission filter or a filter blank.
Record the position of filters to compare later with the plate setup.
7. After the filters or filter blanks have been inserted, replace the rubber plugs over the slots
of the filter wheels.
8. Move the camera housing forward and re-attach the latches.
Fig. 2.2. Installing the filters.
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Emission
Filter Wheel
Slot with Excitation
Filter Wheel
Section 3
Loading Software and Firmware
Installing Software
1. Insert the installation CD into your CD-ROM drive.
2. If the CD does not begin installation automatically, then click on Start and then select
Run.
3. Enter the drive letter for your CD-ROM drive followed by a colon, backslash, "setup"
and click OK. E.g., if your CD-ROM drive is drive D, then you would enter D:\setup and
then click OK.
4. You can respond Next to all prompts except when the installation program prompts for
your name, company and serial number. You must enter something in each of these fields;
we suggest entering the last three digits of the camera serial number in the last field.
Note: If you are installing this over an existing version of the iCycler software you will see
the following prompt.
Click the box that says Don't display this message again and then click Yes.
The installation program will create a folder called Bio-Rad in the Program Files folder
and below that, another folder named iCycler. The executable file iCycler.exe will be placed
in the iCycler folder. You can start the software by using the Windows explorer to find the file
iCycler.exe or you can launch the program from the Start menu.
Several more folders are created inside the iCycler folder: Ini, Emulate, Release and User1.
• The Ini folder contains the iQconfig program used for adjusting the mask and collecting
well factors. It also contains the initial mask and factor files: Mask96.ini and Factor96.ini.
The files TC1.ini and SDD.ini are also present in this folder; these files contain
configuration information for the iCycler program. The iQconfig program requires that the
mask96.ini file be present in the Ini folder before it can run.
• Inside the Emulate folder are three files: Remote.dll, Tcd.dll and Usage.txt. If you want
to emulate protocol execution and data collection, but are not connected to a iQ
Detector, copy these versions of Remote.dll and Tcd.dll into the Windows System fold-
er of your Windows 95 or 98 applications. For Windows NT users, copy these files into
the Winnt System32 folder. This same information is provided in the Usage.txt file.
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• The Release folder contains versions of the Remote.dll and Tcd.dll files that are necessary
for the software to communicate with the iQ Detector. During installation, copies of these
files will be placed in the Windows System folder automatically. If you want to run the
emulation program without a detector, you must first copy the files from the Emulate
folder into the Windows System folder. If you later reconnect the Detector, you must
replace the Remote.dll and Tcd.dll files with the versions in the Release folder. This
information is also provided in the Usage.txt file in the Release folder.
• The User1 folder is where plate setup and protocol files will be saved.
Upgrading the Embedded Firmware. The firmware resident in the iCycler base must be
upgraded to the latest version in order to work properly with the iQ Detector. The upgraded
version of the firmware and a utility for loading it are on the Installation CD-ROM. You must
first create a new folder for the update utility and the new version, copy them from the
CD-ROM to the folder and then run the utility.
1. Connect the 9-pin serial cable from the iCycler serial port to either COM1 or COM2 port
on a Pentium-class computer.
2. Use the Windows Explorer to create a new folder on the C: drive. Select C drive to use
explorer to create a new folder. Name the folder "firmware upgrade".
3. Open the diskette directory \base unit\firmware upgrade and copy the files
UPGRADE.EXE and ICYCUPDT.BIN to the folder \firmware upgrade.
4. Close any other application that could already have taken control of the iCycler base unit.
(ICYCLER.EXE or the report utility software)
5. From the Explorer, open the \firmware upgrade folder and double click on
UPGRADE.EXE. The utility will open and automatically download the new version of
the firmware over the serial port.
6. The UPGRADE utility compares the version of the installed firmware and the version
of the new version provided with the UPGRADE utility.
The screen will show:
Old program version: ____
New program version: ____
Proceed with download? y/n:
Only enter "y" after checking that the new version is a higher number than the old version.
Enter "n" (or any key but "y") if the new version is the same or a lower version number.
7. When the upgrade is complete, a message will be displayed telling you that the upgrade
has been successfully completed. You must turn the iCycler off and back on to
implement the new version of firmware.
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