WPI APOLLO 4000 User manual

APOLLO 4000
World’s first fully-integrated Free Radical Analyzer
INSTRUCTION MANUAL
Serial No._____________________
061506
World Precision Instruments
www.wpiinc.com
APOLLO 4000
WARNING
This instrument must not be connected to a local
network nor to the Internet. Do not attach any
peripheral device other than a USB printer. Any
change to the proprietary hard disk registry in this
device — whether by virus or by normally benign
hardware or software installations — may render the
drive or the Apollo software inoperable, requiring the
instrument’s return to the factory for reformatting.
File corruption or damage to applications or
operating system caused by such use will not be
covered by the Warranty.

APOLLO 4000
WORLD PRECISION INSTRUMENTS 3
Copyright © 2003 by World Precision Instruments, Inc. All rights reserved. No part of this publication may be reproduced or translated into
any language, in any form, without prior written permission of World Precision Instruments, Inc.
Contents
INTRODUCTION ............................................................................... A-1
Design Architecture ........................................................................................ A-2
Plug-and-Play Design ..................................................................................... A-2
Free Radical Sensor Technology .................................................................... A-2
Instrument Description ................................................................................... A-3
Unpacking ...................................................................................................... A-5
OPERATING INSTRUCTIONS ........................................................... B-1
Setting up the APOLLO 4000 ......................................................................... B-1
APOLLO.EXE Operating Software .................................................................. B-4
Example of a real signal application ............................................................ B-12
USING THE APOLLO 4000 TO DETECT NITRIC OXIDE ................... C-1
Initial Set-up .................................................................................................... C-1
Calibration of the NO Sensor .......................................................................... C-1
The Calibration Kit .......................................................................................... C-1
Calibration by the chemical generation of NO ............................................... C-2
Calibration of NO sensor by decomposition of SNAP .................................... C-7
Calibration of NO sensor using aqueous standards prepared with NO Gas C-15
Measurement of NO ..................................................................................... C-18
Maintenance of NO Sensors ........................................................................ C-20
USING THE APOLLO 4000 TO DETECT OXYGEN ............................ D-1
Initial Set-up .................................................................................................... D-1
Calibration and Use of Oxygen Sensors ........................................................ D-1
Calibration ...................................................................................................... D-2
Probe Structure and Assembly....................................................................... D-8
USING THE APOLLO 4000 TO DETECT HYDROGEN PEROXIDE..... E-1
Initial Set-up ..................................................................................................... E-1
The structure of the HPO sensor ..................................................................... E-1
Calibration of the HPO Sensor ........................................................................ E-2
Calibration Procedure ...................................................................................... E-2
Interference ..................................................................................................... E-4
Maintenance of HPO Sensors .........................................................................E-4
TROUBLESHOOTING FOR APOLLO 4000 ........................................ F-1
APPENDIX: BASIC GROUNDING & SHIELDING PRINCIPLES ........ G-1

APOLLO 4000
INTRODUCTION
WORLD PRECISION INSTRUMENTS A-1
INTRODUCTION
The APOLLO 4000 is the end result of an
extensive three-year research and
development program aimed at designing the
most advanced multifunctional free radical
detection system available. Building on
WPI’s worldwide-recognized expertise in
the field of nitric oxide detection and the
success of its popular NO detector (the
ISO-NO series), WPI’s scientists
embarked on an ambitious plan to
develop a state-of-the-art free radical
detection system incorporating the very
latest digital signal processing (DSP) technology.
The APOLLO 4000 is an optically isolated multi-channel electrode-based free
radical analyzer designed specifically for the detection of a variety of redox-
reactive species of biomedical importance. The electrochemical (amperometric)
detection principle used is similar to that employed in WPI’s popular nitric oxide
detection system, the ISO-NO Mark II. However, the APOLLO 4000 incorporates
numerous highly advanced design features that enable it to detect a broad range
of redox-reactive species with unsurpassed accuracy and sensitivity. Using WPI’s
extensive range of free radical sensing electrodes the APOLLO 4000 is able to
detect nitric oxide, hydrogen peroxide, s-nitrosothiols and oxygen. On-going
research at WPI is focusing on expanding the range of detectable species.
NO sensors used with ISO-NO Mark II are completely compatible with the APOLLO
4000.

APOLLO 4000
INTRODUCTION
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Design Architecture
The APOLLO 4000 is based on an optically isolated 4-channel configuration (a 2-
channel version is also available). This design enables simultaneous real-time
measurement of NO (or other free radicals) to be performed using up to 4 different
electrodes. In addition, each free radical sensing channel also contains an
independent channel for temperature measurement.
APOLLO 4000 incorporates a powerful single board computer and proprietary
software (apollo.exe) that enables real-time display and data-acquisition of
individual channels or any combination of channels. An extensive graphical user
interface (GUI) based on a full color LCD monitor allows complete control and
programming of all detection and data-acquisition parameters to be made using
the standard keyboard and mouse included with the system.
The APOLLO 4000 consists of two functionally independent modules: Front End
Converter (FEC) and User Interface (UI). The FEC is an 8-channel data-acquisition
module based on a 24-bit A-to-D and 16-bit D-to-A conversion driven by a Digital
Signal Processor (DSP). The User Interface is built on a standard PC platform with
Windows 2000®operating system. A standard serial port (RS232) provides the
communication between the FEC and UI. The system is fully compatible with a
standard keyboard and mouse and can be readily interfaced with PC’s, computer
networks, printers, and any device that uses Ethernet Tbase-10/100, USB, Serial
Port or Parallel Port communications.
Plug-and-Play Design
The APOLLO 4000 is designed for use with WPI’s range of free radical sensors.
The user simply plugs the required sensor into any one of the input channels
located on the instruments main front panel and then selects the detection and
acquisition parameters using the integrated software control. Each channel is also
provided with an independent temperature input port that allows real-time
monitoring of temperature using WPI’s appropriate temperature sensors.
Free Radical Sensor Technology
The APOLLO 4000 and its associated free radical sensors can provide fast,
accurate, and stable measurements over a wide range of concentrations in both
aqueous solutions and in gas mixtures. Its features include a rapid response time,

APOLLO 4000
INTRODUCTION
WORLD PRECISION INSTRUMENTS A-3
high sensitivity and selectivity, ease of use, and versatility unmatched by any other
similar instrument.
The detection principles are based on the electrochemical (amperometric)
response produced by the various compatible free radical sensors. In summary,
the free radical of interest diffuses through a selective membrane covering the
sensor and is oxidized at the working electrode, resulting in an electrical (redox)
current. The amount of redox current produced is proportional to the free radical
concentration in the sample. All of WPI’s free radical sensors are “combination
electrodes” in which the sensing and reference electrodes have been combined
within a high performance Faraday shield designed to minimize susceptibility to
environmental noise. The Apollo software can be programmed to display either
redox current ( e.g. , pA) or concentration ( e.g. , nM). Output from the APOLLO 4000
can also be collected via BNC connectors on the rear panel of the instrument.
APOLLO 4000 is fully compatible with WPI’s extensive range of free radical
sensors. Currently this range of sensors includes electrodes for monitoring; nitric
oxide, oxygen and hydrogen peroxide. However, new sensors are currently in
development. For details on the complete list of compatible sensors please see the
latest WPI product catalog, or visit WPI’s website (www.wpiinc.com).
Instrument Description
Parts List
The package should contain the following items:
Part No. Description
APOLLO4000 Free Radical Analyzer
800408 Standard Keyboard
800407 Standard Mouse
800406 17” LCD monitor
35209 Program CD
—Power cables
—Instruction Manual

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INTRODUCTION
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}
Channels
Temperature
LED (red)
Sensor
LED
(green)
Power
Temperature
Input
Sensor Input
DVD / CD-RW
Drive
Mains
Power
110V/220V
Switch
mouse
Power
Supply for
Monitor
Keyboard
Analog
Outputs
(BNC)
USB
Ethernet
Serial
Video
Audio
Output
Audio
Input
LR
Parallel Joystick

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INTRODUCTION
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Unpacking
Upon receipt of this product, make a thorough inspection of the contents and check
for possible damage. Missing cartons or obvious damage to cartons should be noted
on the delivery receipt before signing. Concealed loss or damage should be re-
ported at once to the carrier and an inspection requested. Please read the section
entitled “Claims and Returns” on the Warranty page of this manual.
Returns: Do not return any goods to WPI without obtaining prior approval (RMA #
required) and instructions from our Returns Department. Goods returned (unautho-
rized) by collect freight may be refused. If a return shipment is necessary, use the
original container. If the original container is not available, use a suitable substitute
that is rigid and of adequate size. Wrap the instrument in paper or plastic surrounded
with at least 100 mm (four inches) of shock absorbing material. Please read the sec-
tion entitled “Claims and Returns” on the Warranty page of this manual.

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INTRODUCTION
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APOLLO 4000
OPERATING INSTRUCTIONS
WORLD PRECISION INSTRUMENTS B-1
OPERATING INSTRUCTIONS
To exploit the APOLLO 4000’s capabilities fully, it is very important that the user be
aware of the general methods for operating the maintaining the instrument. This will
also ensure that the user is able to understand and interpret the readings.
Setting up the APOLLO 4000
1. Place the APOLLO 4000 on a secure, flat surface ( e.g. , laboratory bench).
2. Position the LCD display on top of the APOLLO 4000 and connect the
display cable to display output video port located on the rear panel of the
APOLLO 4000 (see A-4).
3. Power the LCD display by connecting one end of the cable from the
provided voltage converter (AC to 12V DC) to the LCD display Power input
receptacle. Connect the other end of the voltage converter to a matching
3-prong grounded wall receptacle and switch on.
4. Connect the power supply cord to the back of the APOLLO 4000 and plug
the other end into a matching 3-prong grounded wall receptacle (see set-
up diagram below).
NOTE: Ensure that the red voltage selector switch on the rear panel
(next to the power cable receptacle) is set to the correct voltage —
220 or 110.
Switching ON the APOLLO 4000
The unit can be turned on by pressing the “Power” pushbutton on the front panel.
After booting up, the system automatically starts the main application software
(APOLLO.exe). If other applications have been used the user can return to
Apollo4000 application by double clicking the “Apollo” icon in the right upper
corner of Windows®desktop.
During the process of booting up no error messages should appear on the screen.
(see Troubleshooting).
Switching OFF the APOLLO 4000
The APOLLO 4000 incorporates a highly advanced single board computer and

APOLLO 4000
OPERATING INSTRUCTIONS
WORLD PRECISION INSTRUMENTSB-2
Fig. B1 – When
attaching a
temperature probe
or sensor to the
APOLLO4000,
align the red dot of
the instrument with
the red dot on the
cable connector.
associated electronics. To turn the unit off, it is therefore only necessary to press
the Power button once. It is recommended, however, to close the application
programs
before pressing the Power button. In some cases when the embedded
computer is not responding it may be necessary to press and hold the Power
button in for 3 seconds before the unit will turn off (see troubleshooting section).
The alternative method for switching the unit off is to position the mouse cusor on
START, click on hold down once, and then choose SHUT DOWN from the pop-up
menu. This method will be familiar to Windows®users.
Precautions for handling sensors
The range of free radical sensors offered by WPI vary in their fragility. However, at
all times the user must exercise caution to avoid damaging the delicate
polymeric membrane covering the end of each sensor. This membrane
prevents water and dissolved species such as ions and macromolecules from
reaching the electrode surface where they would interfere with normal
measurement and poison the electrode surface. When the sensor membrane
becomes damaged, sample contents are free to react at the electrode surface.
This causes the background current to become very large and/or go off scale
(negative or positive depending on the reacting species). The membrane integrity
of the sensor can always be checked by ascertaining that the current remains low
and stable when the sensor tip is immersed in a 1.0 M saline solution.
Attaching a sensor to the APOLLO 4000
Each channel on the APOLLO 4000 is equipped with two high quality sensor input
receptacles. The channel marked “Temp” is for use only with a compatible WPI
temperature electrode ( e.g. , ISO-TEMP-2). The channel marked “Sensor” is for use
only with one of WPI’s free radical sensors.

APOLLO 4000
OPERATING INSTRUCTIONS
WORLD PRECISION INSTRUMENTS B-3
NOTE: The Temp and Sensor inputs are not interchangeable but no damage
to the APOLLO 4000 will occur if a sensor/electrode is accidentally inserted
into the wrong input receptacle.
To connect a sensor, simply line up the red dot on the metal connector attached to
the sensor cable with the red dot on the sensor input receptacle and insert the
cable connector (Fig. B1). An LED (located above each sensor input) will light up
immediately indicating the instrument and sensor are connected and working
correctly. The Temp LED is red. The Sensor LED is green. The following LED
indications will inform the user of the status of the system:
LED CONDITION INDICATION
Temp LED (red)
Steady RED light Electrode is performing normally
No RED light Electrode is not connected or is damaged
Sensor LED (green)
Steady GREEN light Sensor is performing normally within the
user-selected current range
Intermittent blinking GREEN light Sensor current is outside the user-
selected linear range.
No GREEN light ERROR sensor current range too low.
Sensor error or sensor not connected.
NOTE: WPI strongly recommends that APOLLO 4000 be powered
through a Back-UPS unit to avoid system failure during power loss.
It is the responsibility of the user as well to install appropriate anti-
virus protection software.
WPI will not be liable for any loss of data as a result of power loss or virus-attack to the APOLLO 4000 system.

APOLLO 4000
OPERATING INSTRUCTIONS
WORLD PRECISION INSTRUMENTSB-4
Fig. B2
APOLLO.EXE Operating Software
The operating software of the APOLLO 4000 is based on a standard Windows®
format, hence many of the software control features will already be familiar to the
user.
Main Screen
The main working screen is
shown in Fig. B2. The software
automatically recalls the last
settings used, therefore the
appearance of the screen will
depend on how it was last used
immediately before being turned
off.
In the example shown, the set-up
is for a four-channel nitric oxide
application with horizonal scale of
6 sec/division and vertical scales
5000 pA/div for channels 1 to 4.
Data for each channel is
displayed on the left of the screen. Each data channel display shows the following
information:
1. Zeroed — Relative measurement value (includes any zeroing applied to
signal).
2. Unzeroed — Absolute measurement value ( i.e. , true value without
zeroing) often refered as “background signal”.
3. Temp — Temperature measurement in degrees Celsius.
4. Unit — e.g., pA, nM, etc.
Note: If no temperature sensor is connected there will be no value in the temp
window and no trace in the graph.

APOLLO 4000
OPERATING INSTRUCTIONS
WORLD PRECISION INSTRUMENTS B-5
Menu System
The Apollo 4000 software uses standard Windows®controls. Help notes will
therefore automatically appear when the cursor is positioned on any control
function or word.
The following section contains a brief description of the programs
main menu system. However, in most cases a user familiar with
Windows®-based operating programs will be able to operate the
software efficiently with the minimum of instructions.
File Menu (Fig. B3) consists of typical Windows®commands:
•New — starts a new data file
•Open — opens an existing data file
•Save — saves the current experiment with the default name
•Save as — allows the user to chose the name and the path of the saving
data file
•Print — prints the screen to the system default printer
•Exit — exits the program and returns to the Windows®desktop.
The standard message (Fig. B4) appears when it is needed. For a
detailed explanation of the above commands please refer to a
Windows®textbook.
Fig. B5 —
Setup Menu
Fig. B3 —
File Menu
Fig. B4
Setup Menu (Fig. B5) consists of the following menu sub-
commands:
•Number of Channels — Sets the desired number of
displayed channels. When One is selected then only the
Channel 1 is displayed. Select Two Channels and 1 and 2
are displayed. Select Three channels for 1, 2, and 3. Select
Four to display all four channels.

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OPERATING INSTRUCTIONS
WORLD PRECISION INSTRUMENTSB-6
•Sample Rate — Selecting this command triggers the
following sequence of events:
Current sampling rate is indicated (Fig. B6). If this
value is satisfactory, the user can chose Cancel and
continue to work with the current sampling rate.
NOTE: The default sampling rate is 10 samples per second, sufficient
for most applications.
To select a different sampling rate, select OK. The sampling rate change
screen (Fig. B7) will appear following OK confirmation by another
confirmation screen (Fig. B8).
The user can cancel the
action and continue to
work with the current
setting. Changing the
sampling rate changes
the data file structure
and it is therefore
necessary to close the
previous file (Fig. B4).
This can interrupt the
current experiment
which may be undesirable. Therefore the user must carefully select the
sampling rate before the experiment. Sampling rate can be set from 1 to
50 samples/sec in increments of 5. It is very important for the user to
understand that Windows®-based computers have the limitations of 16,384
pixels per screen and therefore (number of horizontal divisions=10) the
maximum horizontal scale value will be limited to an integer of 1,638.4/
sample rate. After the new sample rate is selected the horizontal scale
factor is set (for clarification) to the absolute maximum value. For instance,
if the sample rate is set to 5
samples/sec the horizontal
scale will be set to 327 sec/div.
The user can reset it to any
lower value (not lower than 1
sec/div).
Fig. B8
Fig. B6
Fig. B7

APOLLO 4000
OPERATING INSTRUCTIONS
WORLD PRECISION INSTRUMENTS B-7
• Range . After selecting this submenu the range control screen appears
(Fig. B9). The user must select a current range for each sensor channel
within the maximum measured/expected value of the experiment. Proper
selection of the measurement range is very important because the
dynamic range of the instrument is
limited to approximately 1,000,000. This
means that the intrinsic background
noise ( i.e. , “noise floor”) is proportional
to the maximum measured value. For
example:
If 10 nA range is selected, then the
noise floor will be approximately 10 nA
divided by 1,000,000 ( i.e. , 10 fA).
Conversely, if 10 µA range is selected
then the noise floor will be
approximately 10 µA divided by
1,000,000 ( i.e. , 10 pA).
If an incorrect range is chosen for any
channel, the Apollo 4000 will indicate
this as follows:
Fig. B9
Green (sensor) LED Indication Remedy
Intermittent blinking Current detected by sensor is too high Select a higher range
for the selected range
No Green Light Current detected by sensor is too low Select a lower range
for the selected range
Steady Green Light Sensor is normal and within the selected range No action required
If the user is satisfied with the current setting he can chose Cancel and
work as previously.

APOLLO 4000
OPERATING INSTRUCTIONS
WORLD PRECISION INSTRUMENTSB-8
Fig. B11
•Poise Voltage . This submenu looks (Fig. B10) and works similarly to the
Range control. Threre are four choices:
Nitric Oxide —
Automatically configures
the poise voltage ( i.e. ,
865 mV) on the selected
channel to measure nitric
oxide.
Oxygen — Automatically
configures the poise
voltage ( i.e. , 700 mV) on
the selected channel to
measure oxygen.
Hydrogen Peroxide —
Automatically configures
the poise voltage ( i.e. ,
400 mV) on the selected channel to measure
hydrogen peroxide.
Custom — Allows the user to manually set
the poise voltage ( i.e ., from 0-2000 mV)
(Fig. B11). Selecting the wrong poise
voltage will drastically change the results
of an experiment and may render any data
invalid. Hence the program asks for the
confirmation (Fig. B12) prior to changing any
poise voltage.
Fig. B10
Fig. B12

APOLLO 4000
OPERATING INSTRUCTIONS
WORLD PRECISION INSTRUMENTS B-9
•Unit Conversion. There are four default units of measuring signals: fA,
pA, nA and µA. If a new unit needs to be defined chose the Unit
Conversion
submenu in the Setup control. The Amplitude Unit
Conversion screen appears (Fig. B13).
Example: The user defines a new unit called
“Custom” with the conversion ratio 1 unit = 186
pA (Fig.B14). If Add Unit button is selected then
the new unit name appears in the list for the
particular channel (Fig.B15). The user can define
as many custom units as needed as well as
modify the existing custom ones.
Fig. B15
Fig. B13
Fig. B14

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WORLD PRECISION INSTRUMENTSB-10
•On / Off toggle control (also the push button in the
left bottom corner of the screen). This menu starts
and stops the acquisition of data, including writing to
a data file. Before the acquisition starts, the program
notifies the user about the sampling rate and
maximum horizontal scale factor (Fig. B16).
•Plot / Analyze toggle control (also the push button in the left bottom corner
of the screen) commands the program to start and stop plotting of the
incoming data to the screen. In the Analyze mode there is a possibility to
measure different parameters of the acquired data defined by Calculate
menu with the cursors. The cursors appear when the pointer device
(mouse) is moved to a certain position and its left button pushed. There are
a total of two cursors. The second cursor appears after the first one when
the pointer device left button is released .
Calculate Menu includes five different self explanatory parameters that the
program can recalculate from the recorded data stream (Fig.B17):
•Value and Delta — The value as well as time parameter of
plotted data is indicated in the appropriate windows (see
Fig. B2) when the pointer device moves. Clicking of the pointer
device left button fixes the first cursor. While the left button is
depressed the first cursor position data is subtracted from the
data of the current cursor position. When the left button is
released the second cursor is fixed and the difference (delta)
between second and first cursors is measured in the approriate amplitude
and time windows.
•Samples, Average, Min, Max modes enable the appropriate figure
measured between the cursors. It is either number of samples, average,
minimum or maximum value accordingly.
Fig. B16
Fig. B17