Bio-Rad EXQuest Spot Cutter User Manual

PDQuest

™

User Guide for Version 7.1
Windows and Macintosh

P/N 4000129-14 RevA

PDQuest User Guide

Bio-Rad Technical Service Department

Phone: [phone]

510-741-2612
Fax: [phone]
E-mail: [e-mail]

Notice:

No part of this publication may be reproduced or transmitted in any form or by any
means, electronic or mechanical, including photocopy, recording, or an y information
storage or retrieval system, without permission in writing from Bio-Rad.

PDQuest and The Discovery Series are trademarks of Bio-Rad Laboratories. All
other trademarks and registered trademarks are of their respective companies.

WASTE text engine © 1993–1997 Marco Piovanelli

Limitations of Liabi lit y :

Bio-Rad is not responsible for the misinterpretation o f res ults obtained by following
the instructions in this guide. Whenever possible, you should contact the Technical
Services Department at Bio-Rad to discuss your results. As with all scientific studies,
we recommend that you repeat your experiment at least once before making any
significant conclusions for presentation or publication.

Copyright © 2002 by Bio-Rad Laboratories. All rights reserved.

ii

Table of Contents

1. Introduction ........................................................................ 1-1

1.1. Overview of PDQuest ..................................................................................... 1-1

1.2. Digital Data and Signal Intensity .................................................................... 1-2

1.3. PDQuest Workflow ....................................................................................... 1-4

1.4. Computer Requirements .................................. ............................................... 1-7

1.5. Installation ...................................................................................................... 1-9

1.6. Hardware Security Key (HSK) ..................................................................... 1-10

1.7. Starting the Program ..................................................................................... 1-12

1.8. Software License .......................................................................................... 1-13

1.9. Downloading from the Internet .................................................................... 1-16

1.10. PDQuest Basic ............................................................................................ 1-17

1.11. Contacting Bio-Rad ........................................................ ...... ...................... 1-17

2. General Operation .............................................................. 2-1

2.1. Graphical Interface ......................................................................................... 2-1

2.2. Keyboard Shortcuts ........................................................................................ 2-5

2.3. File Commands and Functions ....................................................................... 2-7

2.4. Printing and Exporting .................................................................................. 2-20

2.5. Printing ......................................................................................................... 2-21

2.6. Exporting ...................................................................................................... 2-25

2.7. Preferences .................................................................................................... 2-35

2.8. Mouse-assignable Tools ............................................................................... 2-45

3. Viewing and Editing Images ............................................. 3-1

3.1. Windows and Subwindows ......................................... .................................. .. 3-1

3.2. Configuring Subwindows .......................... ...... ...... .................................. ..... .. 3-1

3.3. Assigning and Interchanging Images .............................................................. 3-3

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PDQuest User Guide

3.4. Tiling Windows ............................................................................................... 3-4

3.5. Magnifying Images .............................................................. ........................... 3-4

3.6. Positioning Images .......................................................................................... 3-7

3.7. Hiding Overlays ............................................................................................ 3-10

3.8. Density Tools ................................................................................................ 3-11

3.9. Colors ............................................................................................................ 3-13

3.10. Multi-Channel Viewer ....................................................... ..... .................... 3-16

3.11. 3D Viewer ................................................................................................... 3-18

3.12. Transform .................................................................................................... 3-20

3.13. Cropping Images ..................................................................... ...... ...... ........ 3-26

3.14. Flipping and Rotating Images ..................................................................... 3-29

3.15. Filtering Images .......................................................................................... 3-32

3.16. Invert Data ................................................................................................... 3-38

3.17. Text Overlays ..................................................................... ..... ...... .............. 3-38

4. Detecting and Editing Spots ............................................. 4-1

4.1. Selecting Spot Detection Parameters .............................................................. 4-1

4.2. Detecting Spots .............................................................................................. 4-9

4.3. Filtered and Gaussian Images ....................................................................... 4-12

4.4. Spot Crosshairs and Ellipses ......................................................................... 4-14

4.5. Adding and Removing Spots ........................................................................ 4-15

4.6. Spot Boundary Tools .................................................................................... 4-18

4.7. Cancelling Spots ........................................................................................... 4-22

4.8. Combining Spots ........................................................................................... 4-23

4.9. Saturated and Faint Spots .............................................................................. 4-23

4.10. Spot Quantity .............................................................................................. 4-24

4.11. Spot Quality ................................................................................................ 4-26

4.12. Spot Parameters ........................................................................................... 4-28

4.13. Finding Spots .............................................................................................. 4-28

5. MatchSets ........................................................................... 5-1

5.1. Creating a MatchSet ........................................................................................ 5-3

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Contents

5.2. Selecting the Master ...................................................................................... 5-9

5.3. Matching Spots ..................................... ..... .................................. ...... ...... ..... 5-10

5.4. Landmarking Spots ....................................................................................... 5-14

5.5. Automated Matching ................................................................................... 5-17

5.6. Cybergels ...................................................................................................... 5-23

5.7. Adding Spots to the Master .......................................................................... 5-26

5.8. Edit Match Submenu .................................................................................... 5-29

5.9. Matching Summary ................................... ...... ...... .................................. ..... 5-37

6. Analysis Tools .................................................................... 6-1

6.1. Spot Review Tool ........................................................................................... 6-1

6.2. Image Stack Tool ............................................................................................ 6-5

6.3. Scatter Plot Tool ............................................................................................. 6-7

6.4. Replicate Groups ............................................................................................ 6-9

6.5. Group Consensus .......................................................................................... 6-13

6.6. Sample Database ........................................................................................... 6-17

6.7. Normalization ............................................................................................... 6-23

6.8. MrpI Data .............................................. ..... .................................. ...... ...... ..... 6-29

6.9. Standard Spot Numbers ................................................................................ 6-32

6.10. Database Spot Numbers .............................................................................. 6-34

7. Analysis Sets and Annotations ........................................ 7-1

7.1. Analysis Sets ................................................................................................... 7-1

7.2. Annotation Tool ............................................................................................ 7-22

7.3. Browsing Annotations .................................................................................. 7-33

7.4. Creating Annotations from Analysis Sets ..................................................... 7-34

7.5. Printing Annotations ..................................................................................... 7-36

7.6. Transferring Annotations ............................................................. ................. 7-37

8. Basic Excision Tool ........................................................... 8-1

8.1. Spot Cutter Setup ............................................................................................ 8-2

8.2. Cutting Spots .................................................................................................. 8-8

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PDQuest User Guide

8.3. Other Spot Cutter Controls .................................................. ..... .................... 8-23

9. Integrated Excision Tool ................................................... 9-1

9.1. Excision Gel Selection .................................................................................... 9-2

9.2. Integrated Excision Tool ................................................................................. 9-8

10. Mass Spectrometry Analysis .......................................... 10-1

10.1. Creating a MassLynx Batch File ................................................................. 10-1

10.2. Importing MassLynx Results ...................................................................... 10-3

10.3. Mass Spec Score Overlay ........................................................................... 10-5

10.4. Protein Probe ..................................................... .......................................... 10-6

10.5. Import Mascot Results ................................................................................ 10-7

11. Graphs and Reports ........................................................ 11-1

11.1. Configure Graphs ........................................................................................ 11-1

11.2. Histogram Graphs ................................................................... ...... ...... ........ 11-4

11.3. A-B Overlay ................................................................................................ 11-7

11.4. Quantity Table Report ................................................................................. 11-9

11.5. Master Image Report ................................................................................. 11-13

11.6. Scatter Plot Report .................................................................................... 11-14

Appendix A.

Gel Doc 2000 ............................................................................. A-1

Appendix B.

ChemiDoc .................................................................................. B-1

Appendix C.

ChemiDoc XRS ......................................................................... C-1

Appendix D.

GS-700 Imaging Densitometer ................................................ D-1

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Contents

Appendix E.

GS-710 Imaging Densitometer ............................. ..... .... ..... ..... E-1

Appendix F.

GS-800 Imaging Densitometer ................................................ F-1

Appendix G.

Fluor-S MultiImager ............................................................. .... G-1

Appendix H.

Fluor-S MAX MultiImager ......................................................... H-1

Appendix I.

Personal Molecular Imager FX ................................................. I-1

Appendix J.
Molecular Imager FX Family (FX Pro, FX Pro Plus and Molecular

FX) .............................................................................................. J-1

Appendix K.

VersaDoc ................................................................................... K-1

Appendix L.

Calibration and Merging .......................................................... L-1

Appendix M.

Cross-Platform File Exchange ........................................... .... M-1

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PDQuest User Guide

viii

Preface

1. About This Document

This user guide is designed to be used as a reference in your everyday use of
PDQuest™. It provides detailed information about the tools and commands of
PDQuest for the Windows and Macintosh platforms. Any platform differences in
procedures and commands are noted in the text.

This guide assumes you have a working knowledge of your computer operating
system and its conventions, including how to use a mouse and standard menus and
commands, and how to open, save, and close files. For help with any of these
techniques, see the documentation that came with your computer.

This guide uses certain text conventions to describe specific commands and
functions.

Example Indicates
File > Open Choosing the Open command under the File menu.

Dragging Positioning the cursor on an object and holding down

.

the left mouse button while you move the mouse.

CTRL+S Holding down the Control key while typing the letter s.

Right-click/
Left-click/
Double-click

Some of the illustrations of menus and dialog boxes found in this manual are taken
from the Windows version of the software, and some are taken from the Macintosh
version. Both versions of a menu or dialog box will be shown only when there is a
significant difference between the two.

Clicking the right mouse button/
Clicking the left mouse button/
Clicking the left mouse button twice.

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PDQuest User Guide

2. Overview of Raw 2-D Gel Electrophoresis

Raw 2-D electrophoresis is a method for separating proteins and nucleic acids in a
sample into a two-dimensional pattern of spots in a gel. It combines the techniques of
isoelectric focusing (IEF) with SDS-polyacrylamide gel electrophoresis (SDS­PAGE). Since these two separation methods rely on independent properties of
proteins—chemical and physical—this procedure can resolve complex biological
samples with a high degree of specificity and accuracy. The resolved proteins and
polypeptide fractions can be then identified by their molecular weights and charges
(as indicated by their locations in the Raw 2-D gel), as well as by their differential
expression in different samples, proximity to other spots, intensity, etc.

Raw 2-D electrophoresis involves two sequential separations of a sample in
perpendicular directions. The IEF dimension is run first, in tube gels or on
immobilized pH gradient (IPG) strips. After focusing, the tube gel or strip is placed
on top of an SDS-PAGE slab gel and electrophoresed. This technique can resolve
thousands of prot ein spo ts in a sin gle sa mple; these pr oteins can then b e visualized by
metabolic radiolabeling or a variety of staining methods.

Proteomics Applications

Proteomics is the study of protein expression and regulation in cells, tissues, and
entire organisms. Sev eral thous and proteins are expressed at any given moment in an
organism; at the cellular level, dozens of proteins may be expressed and regulated in
fractions of a second.

Two-dimensional gel electrophoresis is a a cornerstone in the study of how proteins
are expressed, regulated, and modified throughout living systems. Developed almost
a quarter of a century ago, Raw 2-D gel techno logy remains one of the most po werfu l
techniques for resolvin g comp le x mixt u res of p rot eins . The technology has improved
significantly over the past several years, with the advent of IPG strips and simplified
gel running techniques, large-format and cyber gels that allow for greater pH range
and specificity, and new stains and staining techniques. In addition, mass
spectrometry now allows for peptide mass fingerprinting of very small amounts of
protein isolated from gels.

Using a combination of these techniques, pharmaceutical companies can now use
Raw 2-D technology for high-throughput screening of drug compound candidates
using protein targets; research laboratories can study large-scale changes in protein

x

Preface

expression; and companies and institutions can cross-identify and catalog hundreds of
thousands of protein specie s at the cellular level. This prov ides an ex cellent technique
for the study of differential gene expression under various growth conditions. Since
the expression and regulation of indivi dual pro teins can be detected,Raw 2-D gels are
an indirect way to monitor gene activity. They allow for the investigation of
quantitative as well as qualitative changes in cellular protein expression.

The environmental conditions of a cell can be changed in order to determine optimal
growth conditions as well as monitor the cell’s response to different stresses.
Environmental conditions that can stress the cell include changes in temperature, pH,
and nutrient availability. Some examples of the chemical stresses that can be placed
on a cell include drug and hormone treatments. Since protein structure and function
are the direct result of gene expression, the loss or change of a protein as detected by
a Raw 2-D gel can be extrapolated back to events occurring at the DNA level.

Many questions encountered when genes are inserted and express ed in bacteria, yeast,
and other cell types can be answered with Raw 2-D analysis: Is the cell making the
protein? Is the cell’s progeny making the protein? Is the protein being made but not
secreted? Have mutations occurred?

Medical Applications

Raw 2-D gels can also have important application s in medical research. For example,
this technique can be used to verify the presence of specific protein markers that are
linked to genetic diseases and disease states. Used in conjunction with other tests,
Raw 2-D gels can be part of medical screening procedures associated with mutations
and teratology linked to genetic damage.

Growth factors are being studied for their role in the regulation of cell growth. Raw 2­D gels can be used to evaluate quantitative and qualitative changes in cellular proteins
in response to growth factor stimulation.

Raw 2-D gels allow the visualization of proteins whose expression is altered as the
result of cell transformation, introducing oncogenes into the host genome.
Assessment of phosphorylation, sulfation, or other secondary modifications could
reveal functional protein pathways affected by on cogene expression . This informatio n
could contribute to a better understanding of cell growth and regulation.

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PDQuest User Guide

Sample Experiments

Valuable information can be learned by exposing cells to a set of specific
experimental conditions and subsequently examining their biological response.

A preliminary in vitro ex peri men t that is us eful when b e ginning Raw 2-D gel work i s
to radiolabel a cell’s proteins to steady-state with
these proteins will help to determine the commonly expressed proteins in the cell
under normal conditions.

Subsequently, other amino acids such as 3H-leucine, 3H-proline, 3H-lysine, etc., can
be used to radiolabel the proteins to steady-state. This will label any proteins that do
not contain methionine and were, therefore, not detected in the first experiment, and
will provide preliminary information on the amino acid composition.

Other experiments complementary to Raw 2-D gels include: cell fractionation
procedures, and post-translational mod ifications (phosphorylation, methylation, etc.).
Data from these experiments can be added to a database, accumulating information
about these proteins.

35

S-methionine. Running gels of

3. Bio-Rad Listens

The staff at Bio-Rad are receptive to your suggestions. Many of the new features and
enhancements in this version of PDQuest are a direct result of con vers ations with ou r
customers. Please let us know what you would like to see in the next version of
PDQuest by faxing, calling, or e-mailing our Technical Services staff. You can also
use Solobug (installed with PDQuest) to make software feature requests.

xii

1. Introduction

1.1 Overview of PDQuest

PDQuest is a software package for imaging, analyzing, and databasing Raw 2-D
electrophoresis gels.

The software runs in a Windows or Macintosh environment and has a graphical
interface with standard pull-down menus, toolbars, and keyboard commands.

PDQuest can acquire images of gels usi ng any of several Bio-Rad imaging syst ems.
An image of a gel is captured using the controls in the imaging device window and
displayed on your computer screen. T he scanned gel can then b e cropped, rotated , etc.
using the image editing controls.

Spot Detection, Analysis and Databasing

With Automated Spot Detection and Matching you can select the gels you want to
anlayze, detect spots of interest, create a MatchSet, and match gels all from one
dialog box. The Spot Detect ion Wizard guides you through the process of identifying
and quantifying the spots in the gel image.

After detection, gels in the same experimental series are placed in a MatchSet for
comparison, statistical analysis, and databasing. Histograms allow you to quickly
compare the quantities of the same spot in all the gels in a matchset. Spots can also be
compared qualitatively, organized into user-defined sets for further analysis, and
annotated and databased for easy identification. Spots from differen t experi ment al
series can be organized and compared in high-level matchsets.

PDQuest can be used to simultaneously analyze thousands of spots on hundreds of
gels. Data can be exported to other applications such as spreadsheets for further
analysis.

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PDQuest User Guide

Mass Spec Analysis

PDQuest is part of Bio-Rad’ s ProteomeWorks protein analysis package, and controls
Bio-Rad’s ProteomeWorks Spot Cutter. You can cut spots from gels or membranes
using PDQuest, digest them, and perform advanced protein analysis using
MicroMass’ s mass spectrometry instruments and software. Data from MicroMass can
then be imported back into PDQuest to be included in spot annotations.

And More...

Scan files acquired in PDQuest can be analyzed using other Bio-Rad Discovery
Series software applications, such as Quantity One or Diversity Database. Scans can
be converted into TIFF format for easy compatibility with other applications.

1.2 Digital Data and Signal Intensity

The Bio-Rad imaging devices supported by PDQuest are light and/or radiation
detectors that convert signals from biological samples into digital data. PDQuest then
displays the digital data on your computer screen, in the form of gray scale or color
images.

A data object as displayed on the computer is composed of tiny individual screen
pixels. Each pixel ha s an X an d Y coo rdinate, an d a valu e Z. Th e X and Y c oordi nates
are the pixel’s horizontal and vertical positions on the image, and the Z value is the
signal intensity of the pixel.

1-2

Introduction

Signal intensity of a single pixel

intensity

3-D View

2-D View

Fig. 1-1. Representation of the pixels in two digitally imaged spots in a gel.

For a data object to be visible and quantifiable, the intensity of its clustered pixels
must be higher than the intensity of the pixels that make up the background of the
image. The total intensity of a data object is the sum of the intensities of all the pixels
that make up the object. The mean intensity of a data object is the total intensity
divided by the number of pixels in the object.

The units of signal intensity are Optical Density (O.D.) in the case of the GS-700 an d
GS-710 densitometers, the Gel Doc and ChemiDoc with a white light source, or the
Fluor-S and Fluor-S MAX MultiImagers with white light illumination. Signal
intensity is expressed in counts when using the Personal FX or FX, or in the case of
the Gel Doc, ChemiDoc, Fluor-S, or Fluor-S MAX when using the UV light source.

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PDQuest User Guide

1.3 PDQuest Workflow

Acquire the image

Size and orient the image

Spot identification

Spot comp a r is on and

matching

Data analysis

Spot cutting and mass spec

analysis

Publish results

Fig. 1-2. S teps involved in using PD Quest.

Image Acquisition

PDQuest can acquire images o f gels using Bio-Rad’s densitometers, storage phosphor
imagers, and camera-based imaging systems.

First you open the acquisition window for your imaging device, and capture the
image using the control s. Gel i mages ar e saved on your hard disk, networ k file server,
or removable storage media. The displayed image in PDQuest is ready for analysis.

Image Sizing and Orientation

In this step, you adjust the size and or ientation of t he image by using the cropp ing and
rotating tools on the Image menu.

1-4

Introduction

Sp ot Identifica tion

Now you are ready to identify the spots in the gel. The Spot Detection Wizard
automates the process of selecting the proper spot detection parameters for your gels.
Using the Wizard, you select the parameters, study the results of spot detection, then
adjust the parameters until you have identified all spots of interest in your gels.

When spots are detected in PDQuest, the original gel image is filtered and smoothed
to clarify the spots, then three-dimensional Gaussian spots are created from the
clarified spots. The end result is three separate images: the original unaltered scan (2­D Scan), the filtered and processed scan (Filtered image), and a synthetic image
containing the Gaussian spots (Gaussian image).

2-D Scan Filtered Image Gaussian I mage

Fig. 1-3. Images created during spot detection.

All spot matching and analysis in PDQuest are performed on Gaussian spots.

What Are Gaussian Spots?

Fuzzy, streaked, or overlapping spots in a 2-D gel can be difficult to accurately
distinguish and quantify. Because the image profile of an ideal spot confirms to a
Gaussian curve, PDQuest uses Gaussian modeling to generate spots that can be
precisely identified and quantitated.

A Gaussian spot is a three-dimensional representation of an original scanned spot.
Gaussian curves are fitted to the scanned spot in the X and Y dimensions, and then
additional modeling is performed to create the final Gaussian spot.

Using Gaussian modeling, you can accurately quantitate overlapping spots, spots in
gel streaks, and multiple spots in dense clusters.

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PDQuest User Guide

Matching and Editing

After you have detected the spots in a gel or set of gels, you are ready to create a
matchset. A matchset is composed of the Raw 2-D, Filtered and Gaussian images of
the gel(s) in an experiment.

In a matchset, the protein spots from the different gels are matched to each other and
are included in a synthetic image called a matchset standard. The standard in cludes all
the information about the spots in the matchset.

As you match the spots in a matchset, you will correct the results of autom a tic spot
detection by comparing the Gaussian spots with the original spots in the Filtered
image.

Data Analysis

PDQuest provides a variety of analytical tools to help you determine which spots are
statistically and scientifically meaningful.

You can normalize the spot quantities in different gels for more accurate comparison.
You can define replicate groups of duplicate gels. You can create groups of spots that
are quantitatively, qualitatively, and statistically sig nificant using analysis sets. You
can compare the similarity of gels using scatter plots, and review the quantitation of
individual spots usin g the Sp ot Review Tool.

Many of these tools are interactive with the matchset, so you can study the actual
spots in the images as you review their quantities and other data.

You can create high-level matchsets to compare the results of different experiments.
And the powerful annotation tool allows you to annotate your spots, link to Internet
protein databases or other files, and create HTML pages of spot data.

Spot Cutting and Mass Spec Analysis

PDQuest is part of Bio-Rad’s ProteomeWorks protein analysis package, and controls
Bio-Rad’s ProteomeWorks Spot Cutter. You can cut spots from gels or membranes,
digest them, and perform advanced protein analysis using MicroMass’s mass
spectrometry instruments and software. Data from MicroMass can then be imported
back into PDQuest to be included in spot annotations.

1-6

Introduction

Publish Results

When your analysis is complete, you can print your experimental data or export it to
another system for further analysis.

1.4 Computer Requirements

This software is supporte d on Windows 98, XP, NT 4.0, and 2000, or on a Macintosh
PowerPC.

The computer memory requirements are mainly determined by the file size of the
images you will scan and analyze. High-resolution image files can be very large. For
this reason, we recommend that you archive images on a network file server or high­capacity removable disk.

PC

The following is the recommended system configuration for installing and running
on a PC:

Operating system: Windows 98 SE

Windows NT 4.0 with service pack 6
Windows 2000
Windows XP

Processor: Pentium ≥ 333 MHz
RAM: ≥ 128 MB or bet ter for Gel Doc , ChemiDoc, ChemiDoc

XRS, and VersaDoc systems.

≥ 256 MB or better for Molecular Imager FX systems,

Personal FX system, and GS-800 densitometer.

Hard disk space: ≥ 3 GB
Monitor: 17" monitor, 1024 x 768 resolution (absolutely required),

True color.

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PDQuest User Guide

SCSI: Required for all Bio-Rad imaging devices except the Gel

Doc, ChemiDoc, ChemiDoc XRS, and VersaDoc systems.
Adaptec SCSI card recommended.

Printer: Optional.

Macintosh

The following is the recommended system configuration for installing and running
on a Macintosh:

Operating system: System 9.0 or higher, excluding Mac OS X.
Processor/Model: PowerPC G3 processor or better.
RAM: ≥ 256 MB for all Bio-Rad imaging systems.
Hard disk space: ≥ 3 GB
Monitor: 17" monitor, 1024 x 768 resolution (absolutely required),

Millions of colors.

SCSI: Required for all Bio-Rad imaging devices except the Gel

Doc, ChemiDoc, ChemiDoc XRS, and VersaDoc systems.
Adaptec SCSI card recommended.

Printer: Optional.

Note: The default amount o f memory assigned to th is program on the Macintosh is 1 28

MB. If the total RAM in your Macintosh is 128 MB or less, you should reduce
the amount of memory assigned to the program to 10 MB
RAM. With the application icon selected, go to File > Get Info in your Finder to
reduce the memory requirements for the application. See your Macintosh
computer documentation for details.

1-8

less than your total

Introduction

1.5 Installation

Windows

Note: Windows NT and 2000 users: You must be a member of the Administrators

group to install Discovery Series software. After installation, members of the

Users group must have “write” access to the Discovery Series folder to use the

software.

Insert the Discovery Series CD-ROM into your computer. The installer will start
automatically. (If the CD does not auto-start, use Windows Explorer to open the root
directory on the CD-ROM and double-click on the Setup.exe file.)

The installer program will guide you through the installation. The installer will create
a default directory under Program Files on your computer called Bio-Rad\The
Discovery Series (you can select your own directory if you wish). The application
program will be placed in the Bin folder inside the Discovery Series folder.
Additional directories for storing user profiles and sample images will also be created

The installer will place an application icon on your desktop and create a Discovery
Series folder under Programs on your Windows Start menu.

After installation, you must reboot your computer before using an imaging device.

Macintosh

Insert the Discovery Series CD-ROM into your Macintosh. The TDS-Mac folder will
open on your desktop, displaying the installers for the Discovery Series applications.
Double-click on the installer for your application.

Fig. 1-4. Installation program icon (Macintosh).

The installer will guide you through a series of screens. The installer will create a
folder on your hard drive that contains the main application and associated sample
images (you can select a different folder if you wish). The installation will also create

1-9

PDQuest User Guide

a folder called The Discovery Series in the Preferences folder in your System Folder;
this contains the Help file and various system files.

Once installation is complete, the folder containing the application icon will appear
open on your desktop.

Before running the software, you must install the Hardware Security K ey on your
computer (unless you have installed a computer download version of the software).

1.6 Hardware Security Key (HSK)

Note: Initial installation of a network server does require the Hardware Security Key

included in the software package. Installation of an additional Network Client
User to a Network License Server System does not require an HSK. Please refer
to the Network License Installation Guide that ships with Network Licenses .

Discovery Series software is password-protected using a Hardware Security Key
(HSK), which is included in your software package. You must attach the Hardware
Security Key to your computer before you can run the software.

Windows

Fig. 1-5. PC Hardware Security Key

Before proceeding, please turn off your computer.
The HSK attaches to the parallel port on the back of your PC. If a printer cable is

attached to this port, turn off the printer and disconnect it. After you have attached the
HSK, you can attach the printer cable to the key itself and restart your computer and
printer.

1-10

Introduction

Note: Some parallel port devices such as zip drives may be incompatible with HSKs.

Please check with your peripherals vendor.

The code for the PC hardware security key is EYYCY. This is printed on the key
itself.

You will also need to install the system driver that allows the computer to recognize
the HSK.

Note: Windows NT and 2000 users must be in the local administrator group to install

the HSK driver.

To install the driver, open the Windows Start menu and select Programs > The
Discovery Series. Select Install HASP Hardware Security Key driver to begin
installation.

Note: Windo ws 98 users must reboot their compu ter after instal ling the HSK. W indows

NT and 2000 users do not have to reboot.

Macintosh

Fig. 1-6. Macintosh Hardware Security Key

Before proceeding, please turn off your Macintosh.
The Macintosh HSK must be inserted in the Apple Desktop Bus (ADB) path. The

ADB port is located on the back of your Macintosh.

Fig. 1-7. Apple Desktop Bus icon on back of Macintosh.

1-11

PDQuest User Guide

The HSK can be inserted at any point in the ADB path—between the computer and
the keyboard, between the keyboard and the mouse, between the keyboard and the
monitor, etc. After you have attached the HSK, you can restart your computer.

The code for the Macintosh HSK is QCDIY. This code is printed on the key itself.

1.7 Starting the Program

The Hardware Security Key must be attached to the co mputer b efore yo u can st art the
software (unless you are using a network license).

Windows

The installation program creates an application icon on your desktop. To start the
program, double-click on this icon.

Fig. 1-8. Application icon.

You can also start the program from the Windows Start menu. Click on the Start
button, select Programs, select The Discovery Series, an d s elect the ap plication n ame.

Macintosh

After installation, the main application folder will be open on your desktop. To start
the program, double-click on the application icon shortcut inside the folder. You can
move this shortcut icon to your desktop.

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Introduction

1.8 Software License

When the software opens for the first time, you will see a Software License screen
that shows the current status of your software license.

With a new HSK or network license, you receive a 30-day temporary license (“Your
license will expire on _______”). The temporary license is designed to give you time
to purchase the software, if you have not already done so.

Fig. 1-9. Temporary license screen.

During the 30-day period, the Software License screen will appear every time you
open the software. To use the software during this period, click on the Run button.

Network license holders can click on the Check License but ton at any t ime durin g the
30-day period to activate their full network license. (If your network license is not
activated when you click on Check License, notify your network administrator.)

HSK users have 30 days to p urchase the s oftwar e and o btain a purchase order number
and software serial number from Bio-Rad. When you have this information, click on
the Check License or Registration Form button in the Software License screen to
register your software.

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PDQuest User Guide

Fig. 1-10. Software License Registration Form.

Fill out the information in the Software License Registration Form. Be sure to enter
your purchase order number and software serial number under the Purchase
Information tab when registering.

Registering by Internet

If you have Internet access from your computer, click on the Submit via Internet
button to send the Software Registration Form directly to Bio-Ra d.

Your information will b e submitted , and a temporary password will be generated
automatically and sent back to you r com puter. Simply continue to run the application
as before.

Bio-Rad will confirm your purchase information and generate a permanent license.
After 2–3 days, click on Check License in the Software License screen again to
update to a permanent password. (The Software License screen will not appear
automatically after the temporary password has been generated; the software will

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Introduction

simply open normally. Go to the Help menu and select Register to open the Software
License screen.)

Registering by Fax or E-mail

If you do not have Internet access, click on the Print button in the Software License
Registration Form and fax the form to Bio-Rad at the number listed on the form.
Alternatively , yo u can en ter the conten ts of the form into an e-mail and send it to Bio­Rad at the address listed in the Registration Form.

Bio-Rad will contact you by fax or e-mail in 2–3 days with a full license.

Entering a P assword

If you fax or e-mail your registration information, you will receive a password from
Bio-Rad. You must enter this password manually.

To enter your password, click on Enter Password in the Software License screen. If
you are not currently in the Software License screen, select Register from the Help
menu.

Fig. 1-11. Enter Password screen.

In the Enter Password screen, type in your password in the field.

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PDQuest User Guide

Once you have typed in the correct passwor d, the OK l ight nex t to th e passwor d fiel d
will change to green and the Enter button will activate. Click on Enter to run the
program.

1.9 Downloading from the Internet

You can download a trial version of the software from Bio-Rad’s Web site. Go to the
Discovery Series download page at www.bio-rad.com and select from the list of
applications. Follow the instructions to download the installer onto your computer,
then run the installer.

After installation, double-click on the application icon to run the program. The
software will open and the Software License screen will be displayed.

Note: If you attempt to start the downloaded program and receive an “Unable to obtain

authorization” message, you will need a Hardware Security Key to run the
program. Contact Bio-Rad to obtain a key.

Fig. 1-12. Free Trial screen.

In the Software License screen, click on the Free Trial button. This will open the
Software License Registration Form. Enter the required information (you will not

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Introduction

have a purchase order number or software serial number, and can leave these fields
blank) and click on Submit Via Internet.

A free trial password will be automatically downloaded to your computer. This
password will allow you to use the software for 30 days.

If you decide to purchase the software during that period, contact Bio-Rad to receive
a software package and a Hardware Security Key. You can then complete the
registration process as described in the previous sections.

1.10 PDQuest Basic

In order to meet requests for additional software copies at low cost when buying an
imaging system, Bio-Rad has provided a scaled down version of PDQuest called
PDQuest Basic. PDQuest Basic looks and acts exactly like the full version of
PDQuest but with the advanced functions inactive (grayed out).

The Basic version of PDQuest includes the following active functionality: Image
acquisition, Transform, Crop, Flip, Rotate, Text Tool, Basic Excision, Print, and
Export to TIFF and Save. PDQuest Basic also allows you to view previously saved
data created using advanced functionality.

PDQuest Basic does not require a software license. To activate the advanced
commands of PDQuest, contact Bio-Rad for a valid user license.

1.11 Contacting Bio-Rad

Bio-Rad technical service hours are from 8:00 a.m. to 4:00 p.m., Pacific Standard
Time in the U.S.

Phone: [phone]

510-741-2612
Fax: [phone]
E-mail: [e-mail]
For software registration:

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