YSI 5300A Operation Manual
YSI 5300A
Biologic al O xyg en
Monitor System
Operations
Manual
Table of Contents
1. Introduction.............................................................................................................. 1
2. Installing and Initial Setup ...................................................................................... 2
2.1 Unpacking the System........................................................................................................2
2.2 Completing the Warranty Card...........................................................................................2
2.3 Viewing the Front and Back Panels ...................................................................................3
2.4 Setting Up the Software......................................................................................................5
2.5 Setting Up the Standard System.........................................................................................7
2.6 Setting Up the Micro System..............................................................................................9
3. Preparing your Bath Assembly............................................................................. 11
3.1 Choosing your Bath Assembly.........................................................................................11
3.2 Setting up your Bath Assembly........................................................................................13
4. Preparing your Oxygen Probe..............................................................................15
4.1 Standard Oxygen Probe....................................................................................................15
4.2 Micro Oxygen Probe ........................................................................................................17
5. Operating your Oxygen Probe.............................................................................. 18
5.1 Principles of Operation.....................................................................................................18
5.2 Electrical Characteristics..................................................................................................18
5.3 Voltage Plateau.................................................................................................................18
5.4 Noise.................................................................................................................................19
5.5 Temperature......................................................................................................................19
5.6 Oxygen Consumption by the Probe..................................................................................19
5.7 Electrolyte Saturation and Bubbles in Electrolyte............................................................20
5.8 Drift of Calibration...........................................................................................................20
5.9 Probe Care and Maintenance............................................................................................20
6. Operating your Oxygen System ........................................................................... 22
6.1 Calibrating........................................................................................................................22
6.2 Daily Probe Test...............................................................................................................24
6.3 Sample Monitoring...........................................................................................................24
7. Troubleshooting.....................................................................................................26
8. Warranty and Repair..............................................................................................28
8.1 Cleaning Instructions........................................................................................................29
8.2 Packing Instructions .........................................................................................................30
9. Required Notice ..................................................................................................... 31
10. Accessories and Replacement Parts................................................................ 32
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11. Appendix A - Specifications..............................................................................33
12. Appendix B – Tables.......................................................................................... 35
13. Appendix C - Required Notice........................................................................... 40
14. Appendix D: Application Notes........................................................................ 41
15. Appendix E: Gas Solubility Relationships...................................................... 42
16. Appendix F: Interfering Gases......................................................................... 43
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1. Introduction
The YSI 5300A Biological Oxygen Monitor is designed to measure oxygen uptake and evolution
in biological systems. It utilizes Clark type polarographic oxygen probes immersed in
magnetically stirred sample chambers, and produces oxygen uptake or evolution curves in 2 to
15 minutes. This two-channel instrument offers simultaneous display, recorder outputs, and RS232 serial outputs for monitoring the results of each evaluation of samples.
Two different sensor systems are available. The "Standard System", for the measurement of
samples of 3 to 8 ml, consists of a 5300A Monitor, a 5301B Standard Bath, two 5331A Oxygen
Probes, and other supplies. The 5304 Micro Adapter Kit will modify the 5301B for use with 1 ml
samples. The "Micro System," for the measurement of a fixed volume sample of 600 µL or for a
continuous flowing sample, includes a 5300A Monitor, one or two 5356 Micro Oxygen
Chambers, one or two 5357 Micro Oxygen Probes, and other supplies. The Micro Oxygen
Chamber and the Micro Probe are manufactured by Instech Laboratories, Inc..
The YSI 5331A and YSI 5357 Oxygen Probes are a complete polarographic system consisting of
a platinum cathode, silver anode and KCl solution held captive around electrodes by a Teflon
membrane fastened with an o-ring. When a polarizing voltage is applied across the probe, all
oxygen in the probe is consumed (reduced) at the cathode and current flows in direct
stoichiometric relation to the rate of oxygen consumption (reduction). Oxygen then diffuses
through the membrane at a rate proportional to the oxygen pressure outside the probe, since
oxygen pressure in the probe is near zero. When steady state conditions are reached in about 30
seconds, current flows through the probe at a rate in proportion to the external oxygen pressure.
The YSI 5301B Bath Assembly has three sample chambers with built-in magnetic stirrers
surrounded by a water bath. The YSI 5301B is able to measure sample size from 3 to 8 mL with
the YSI 5331A probes. The YSI 5304 Micro Adapter Kit modifies the YSI 5301B for use with
1-mL samples and requires the YSI 5331 probes as well. The YSI 5301B Bath Assembly is
connected with a constant temperature circulator (YSI 5310) and together they provide a
temperature controlled environment where the oxygen uptake and evolution can be accurately
monitored in a sample. The Lucite plunger included with the YSI5301 Bath Assembly provides
an access slot for the introduction of samples, inhibitors and activators into the sample chamber.
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2. Installing and Initial Setup
Except when operating at ambient temperatures, a constant temperature circulator is necessary to
control the temperature of the sample chambers (YSI recommends a constant temperature
circulator with a +/- 0.02 °C control for maximum accuracy). A 2 channel chart recorder, serial
printer, or computer (equipped with an RS232 port or data acquisition system) is necessary for a
hard copy of experimental results. Your YSI dealer can recommend and offer suitable circulators
(YSI 5310) and chart recorders (YSI 5320).
2.1 Unpacking the System
When you unpack your new YSI 5300A Biological Oxygen Monitor for the first time, check the
packing list to make sure you have received everything you should have. If there is anything
missing or damaged, call the dealer from whom you purchased the system. If you do not know
which of our authorized dealers sold the system to you, call YSI Customer Service at 800-6598895 or 937-767-7241, or email us at lifesciences@ysi.com and we'll be happy to help you.
2.2 Completing the Warranty Card
Please complete the Warranty Card and return it to YSI. This will record your purchase of this
instrument in our computer system. Once your purchase is recorded, you will receive prompt,
efficient service in the event any part of your YSI 5300A should ever need repair.
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2.3 Viewing the Front and Back Panels
2.3.1 Front Panel
The front panel of the YSI 5300A contains the display screen, TEST/AIR/ O2/ TEST setting
dials, calibration adjustment dial, Menu, and toggling keys as shown below.
Channel 1 Setting Dial
TEST/AIR/O2/TEST
Channel 1
Calibration Dial
Display Screen
Menu Selection Key
Toggling
Up/Down Keys
Channel 2 Setting Dial
TEST/AIR/O2/TEST
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Channel 2
Calibration Dial
2.3.2 Rear Panel
The rear panel contains the connections for the power supply, power supply switch, YSI 5331A
probes, YSI 5357 Micro Oxygen Probes, RS232 serial port, and recorder outputs.
Power
Supply
Connection
RS-232
Serial Port
Connection
Right Channel
YSI 5331A
Oxygen
Probe Input
Right Channel
YSI 5357 Micro
Probe Input
Left Channel
YSI 5357 Micro
Probe Input
Left Channel
YSI 5331A
Oxygen
Probe Input
Power Supply
Switch
Right Channel
Positive (+)
Recorder Output
Right Channel
Negative (-) Ground
Recorder Output
Left Channel
Positive (+)
Recorder Output
Left Channel
Negative (-) Ground
Recorder Output
Power Supply Connection
The power supply connection requires a 6 VDC power supply with a least 200mA current.
Oxygen Probe Connections
The YSI 5331A has a 0.25” Stereo Phone Plug. The YSI 5357/ 5358 Micro Probe System has a
0.173" miniature telephone plug.
Note: The YSI 5331 Oxygen Probes are not compatible with the YSI 5300A Monitoring
System. Only the YSI 5331A oxygen probe and the YSI 5357/ 5358 micro oxygen probe
system are compatible with the YSI 5300A Monitor.
RS232 Connection
The RS232 connection is a standard DB9 connector. Use a straight serial cable (not a null
modem) to connect the YSI 5300A to a computer serial port or a serial printer. See Accessories
and Replacement Parts, Sec. 11 for the YSI item number.
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2.4 Setting Up the Software
The YSI 5300 has two primary modes: monitoring/calibration mode and the zero calibration/
RS-232 setup mode.
1. The monitoring/ calibration mode is the default mode and is shown below. In this mode the
analyzer is constantly measuring oxygen. The analyzer can also be calibrated in this mode
by unlocking the calibration dial and setting the analyzer to the appropriate calibration value.
(See Sec. 6 for calibration instructions.)
%AIR
%AIR
89.0
Press the up and
down ke ys to zero
adjust the reading
89.0
2. To adjust the contrast on the display screen, press the [▲] up soft-key or the [▼] down softkey in the sampling/ calibration mode.
3. To enter the zero calibration/ RS-232 setup mode press the [MENU] soft-key.
4. The Left Channel Zero Calibration screen will first be displayed. A zero percent sample will
be used to adjust at the zero point calibration at this menu. Adjust the zero point to reflect
0.0% with this zero percent oxygen sample. The left channel zero calibration will display
actual oxygen readings. (See Sec 6.1 for zero calibration instructions.)
Left Channel Zero Calibration
0.0
Press MENU to conti nue
5. Press the [MENU] soft-key again and following screen will be displayed.
%AIR
Right Channel Zero Calibration
Press the up and
down ke ys to zero
adjust the reading
Press MENU to conti nue
A zero percent sample will be used to adjust at the zero point calibration at this menu. Adjust the
zero point to reflect 0.0 % with this zero percent oxygen sample. The left channel zero
calibration will display actual oxygen readings. (See Sec 6.1 for zero calibration instructions.)
6. Press the [MENU] soft-key again and following screen will be displayed
0.0
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%AIR
RS232 Serial Output
OFF
Press UP or DOWN to select
--
> ON
Press MENU to continue
7. Select On or Off to activate or disable the RS232 Serial Output option for serial port data
acquisition and collection.
8. Press the [MENU] soft-key again and following screen will be displayed.
Serial Output Data Ra te
10 seco nds
Press UP or DOWN to select
Press MENU to continue
9. Select the serial output data rate via the RS-232 serial output for your data collection needs.
(Output data rate ranges from 1 to 60 seconds). Compare your computer or serial printer’s
requirements with YSI 5300A setting.
10. Press the [MENU] soft-key again and following screen will be displayed.
Serial O utput Baud Rate
2400
4800
Press UP or DOWN to select
--
> 9600
Press MENU to continue
11. Select the appropriate serial output baud rate for your data acquisition requirements.
Compare your computer or serial printer’s requirements with YSI 5300A setting.
12. Press the [MENU] soft-key again and following screen will be displayed.
Serial Output Parity
ODD
EVEN
Press UP or DOWN to select
--
> NONE
Press MENU to continue
13. Select the appropriate serial output parity per your data acquisition requirements. Compare
your computer or serial printer’s requirements with YSI 5300A setting.
14. Press the [MENU] soft-key again and following screen will be displayed.
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Serial O utput Line T erminator
CR
LF
LF/CR
Press UP or DOWN to select
--
> CR/LF
Press MENU to continue
15. Select the appropriate serial output line terminator per your data acquisition requirements.
Compare your computer or serial printer’s requirements with YSI 5300A setting.
16. Press the [MENU] soft-key again and following screen will be displayed.
Probe Bias Voltages
Normal Test
0.70 volts 0.80 volts
Press UP or DOWN to select
Press MENU to continue
17. The probe bias voltages screen shows the difference in the voltages between the normal
mode and the test mode for both probes.
18. Press the [MENU] soft-key again and following screen will be displayed.
Reset All Sys tem Parameters to Default
YES
Select YES and press menu to reset
Press NO and press menu to continue
--
> NO
The Reset All System Parameters to Default screen allows the user to reset the analyzer to the
factory default settings.
NOTE: Inactivity in the zero calibration/ RS-232 setup mode will cause the analyzer to
default to the calibration/ sampling mode after ~15 seconds.
2.5 Setting Up the Standard System
Description
The YSI 5300A Biological Oxygen Monitor standard system consists of the following separate
YSI products:
• 1 YSI 5300A Biological Oxygen Monitor
• 1 YSI 5301B Standard Bath Assembly (includes 1 5350 Membrane Mounting Kit, 1
5225 Sample Chamber kit, tubing, etc.)
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• 2 YSI 5331 Oxygen Probes, each with a 5775 Standard Membrane/KCl Kit
The assembled 5301B Standard Bath Assembly has three sample chambers with magnetic
stirrers, surrounded by a water chamber. A constant temperature circulator is connected to the
bath. Two 5331 Oxygen Probes are prepared for operation using the 5350 Membrane Mounting
Kit to install the parts found in the 5775 Standard Membrane/KCl Kit. The probes are held in
plungers that fit closely into the sample chambers. The probes plug into the back of the
instrument.
Assembling the Standard System:
1. Insert three glass sample chambers from the box marked 5215 (included with the 5301B)
into the holes in the bath cover.
2. Wet three of the large rubber o-rings with distilled or de-ionized water.
3. Place one rubber o-ring over each of the sample chambers near its top (the o-rings should
seat against the top cover).
4. Slide each sample chamber into the bath until it rests on its stop.
5. Place a white nylon hold down ring over each chamber (above the o-ring), push it down
and twist clockwise to secure it under the stainless steel shoulder screws (this will seat the
o-rings against the top cover for a water-tight seal).
Caution: Over-tightening the hold down ring may break the sample chamber.
6. Make sure that the power switches on the 5301B Bath Assembly and 5300A Monitor are
in the off (0) position, then plug the power cords into a properly grounded power
receptacle.
7. If the constant temperature circulator is used for your application, connect the 5301B Bath
Assembly to the circulator using the YSI 5315 Plumbing Kit. (Wetting the tubing ends
with water or alcohol will ease the tubing installation). Though it is not critical which
5301B tubing fitting is used as the inlet, YSI recommends using the bottom fitting.
8. Fill the circulator with distilled water and set the controls to the desired temperature.
Operate the circulator according to its instruction manual. Note: YSI recommends using
only distilled water in the bath. If distilled water is not used, minor rust stains may appear
in the 5301B Bath chamber. They may be removed using a mildly abrasive pad.
9. If the chart recorder is used for your application, connect a recorder to the recorder output
port on the back of the 5300A Monitor. The 5300A Monitor has a self-zeroing feature
with no probes plugged in; the recorder zero reference can be set with this zero output.
NOTE: The red port on the 5300A Monitor back panel is positive and the black port is
negative.
10. If the RS-232 serial cable is used for your application, connect the RS-232 serial cable to
both the YSI 5300A RS-232 port and the acquisition serial data port.
11. Prepare the 5331 Oxygen Probes for operation as described in section 4.1.
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12. Plug the 5331 Oxygen Probes into the jack marked Input on the rear of the 5300A
Monitor.
NOTE: When a 5331 Oxygen Probe and a Micro Oxygen Probe are both plugged into
the same channel’s input jacks, the display and output values reflect the YSI 5357
Micro Oxygen Probe response.
13. Numbered marking tape is supplied with the 5301B Bath Assembly to identify each probe
if desired.
2.6 Setting Up the Micro System
Description
The YSI 5300 Biological Oxygen Monitor Micro System consists of the following components:
• 1 YSI 5300A Biological Oxygen Monitor
• 1 or 2 YSI 5356 Micro Oxygen Chambers
• 1 or 2 YSI 5357 Micro Oxygen Probes
• 1 or 2 YSI 5358 Cable Assemblies
• 1 YSI 5775 Standard Membrane Kit
You can use the 5356 Micro Oxygen Chamber for fixed-volume single samples or for
continuous, flow-through monitoring. You should use a constant temperature circulator to
control the block temperature. A remote magnetic stirrer controller keeps the sample chamber
stirred when you measure fixed-volume samples.
Assembling the Micro System as follows:
1. Assemble the YSI 5356 Micro Oxygen Chamber as described in the Instech Laboratories
Model 600A Instruction Manual provided.
2. Connect the Oxygen Chamber to a constant temperature circulator (tubing not supplied).
3. Fill the circulator with distilled or de-ionized water and set it to the desired temperature.
Operate the circulator according to its instruction manual.
4. If the chart recorder is used for your application, connect a recorder to the recorder ports
on the rear of the monitor. NOTE: The red port on the 5300A Monitor back panel is
positive and the black port is negative.
5. If the RS-232 serial cable is used for your application, connect the RS-232 serial cable to
both the YSI 5300A RS-232 port and the acquisition serial data port.
6. Prepare the Micro Probe for operation as described in the Instech Laboratories Model
125/05 manual.
7. Thread the probe into the 5358 Cable Assembly.
8. With the monitor off, plug the Cable Assembly into the appropriate jack on the back of the
YSI 5300A.
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NOTE: When a Standard Oxygen Probe and a Micro Oxygen Probe are both plugged into the
same channel's input jacks, the display and recorder values reflect the Micro Probe's response.
9. Immerse the probe in distilled water up to 1" above the rubber o-ring until you are ready
to insert it into the sample chamber.
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3. Preparing your Bath Assembly
3.1 Choosing your Bath Assembly
Three bath assemblies are available with for use with the YSI 5300A Biological Oxygen
Monitor. Each is unique and should be considered for the user’s particular need.
3.1.1 YSI 5301B Bath Assembly
The YSI 5301B Bath Assembly (See Figure 3.1) provides relatively airtight sample chambers
that are stirred magnetically and controlled in temperature when connected to a constant
temperature circulator. The YSI 5301B Bath Assembly is designed to test sample sizes ranging
from 3 to 8 milliliters in volume.
Figure 3.1 YSI 5301B Sample Chamber
The three sample chambers are held in place with locking nylon hold down rings and sealed in
the bath with two rubber o-rings (See Figure 3.2).
The nylon hold down ring also serves as a tensioning device to hold the Lucite plunger in
position while allowing a very fine adjustment regardless of sample size.
The white Lucite plunger has a slanting face and an access slot along the side for removal of gas
bubbles from the sample. The removal of bubbles is of the greatest importance because of the 20
to 1 increase in the amount of oxygen per unit volume in the gas phase over that in saturated
solution.
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