When starting AutoResp™ and this error message appear, AutoResp™ was not installed with administrator rights. During the AutoResp™ installation process, a file must be copied into a Windows system folder. This requires an administrator login. Here is the instruction how to install the necessary file to get rid of the error message:
If the display on your OXY-REG is giving the error message "SE.BR" (sensor breakage), it means that the signal input type for the instrument has been changed from the default setting (potmeter).
To change the input type back to the default setting, press the OK button once. Then press the arrow buttons until the input type POTM (potmeter) is shown in the display and then press OK to choose this. Finally exit the set up menu by pressing the OK button several times until the display shows - - - -.
Tracking requires two synchronized (in time) cameras to track animals in three dimensions.
Mount the two cameras with a 90° angle in between, e.g. one camera filming from above and one filming from the side.
Simultaneous video recordings from both cameras can then be analyzed in LoliTrack to get two pairs of X-Y coordinates.
In this way, one pair of X-Y coordinates translates into the third (Z) coordinate. See the figure below.
This guide shows you how to draw zones and mask in LoliTrack and ShuttleSoft.
Use the Circle button, then draw a circle in one of the compartments. Afterwards, click on the Mask Inside Area button.
Use the Circle button again to draw a circle in the other compartment. Afterwards, click on the Mask Inside Area button.
Use the Rectangular button, then draw a rectangle between the two compartments. Afterwards, click on the Mask Inside Area button.
Now invert the mask by clicking on the Invert Mask button.
Draw a rectangular area in one compartment, and click on a Zone button (A, B, C...). This area is now defined as a zone of interest.
Draw a rectangular area in the other compartment, and click on a different Zone button. This area is now also defined as a zone of interest.
This is an example of how a typical mask and zones could look like running a ShuttleSoft experiment.Link
To change the language in the PIT tag reader software (APR-PC-DEMO) from German to English, please go to the following program files folder on your PC:
Loligo® Systems offers the three main types of oxygen sensors differing in measuring principle, response time, sensor size, maintenance requirement and pricing.
1. Optical oxygen sensors
For many invasive techniques, measurements in tiny volumes (e.g. micro respirometry) or applications which require high temporal and spatial resolution, fibre-optic oxygen sensors are the only solution.
Even for general purposes we recommend optical oxygen sensors featuring low maintenance requirements, high stability and accuracy, no electrical interference, no ground loop problems, and zero O2 consumption by the sensor.
The main disadvantages is price, single-channel meters start at about €3750. A high temperature sensitivity of this technology and a fragile tip on the <50-140 nm micro sensors might also be a problem in some applications. The macro sensors, on the other hand, are very tough.
2. Galvanic cell oxygen electrodes
Galvanic type oxygen probes are inexpensive and rugged sensors producing a milliVolt signal for easy instrumentation without supplying power.
We recommend galvanic probes for applications like respirometry and field measurements, and as a low-budget alternative to optical oxygen equipment, e.g. for multi-channel systems. They can be used with relatively long cables (+25 meters). We recommend using a galvanic isolation preamplifier between the probe and any data acquisition system to minimize possible ground loop problems.
The main disadvantage of galvanic probes is a relative long response time and oxygen self-consumption making them unsuitable for measurements in small volumes and in un-mixed samples.
3. Polarographic oxygen electrodes (or Clark type electrodes)
Low oxygen self-consumption and a small size make these sensors suitable for physiological setups like blood gas analysis and low volume respirometry.
However, Clark type electrodes require much maintenance, often membranes and electrolyte fluid should be changed on a daily basis, and polarographic amplifiers for these sensors are quite costly.
Thus, we recommend the E101 as a replacement oxygen electrode for customers with their own polarograhic oxygen meter, e.g. PHM meter from Radiometer Copenhagen, OM200 from Cameron Instr. Co. etc.Link
Instrument calibration is an essential first step in analytical and measurement procedures. The rationale for a two-point calibration is to provide two points of reference with which to calibrate the instrument and, therefore, ensuring accurate measurements with your sensor of the unknown concentrations of your samples. A two-point calibration between two extremes are advocated, e.g. 0 - 100 % air saturation. For the low calibration point, we recommend using either nitrogen gas or sodium sulphite solution, for the high calibration point, bubble atmospheric air using an aerator. Please see below for general step-wise instructions on how to carry out a two point calibration. For specific instructions for a particular oxygen instrument, please click on the specific links.
See below for a diagrammatic representation of sensor readings at:
|100% to 0% air saturation||0% to 100% air saturation|
|100% to 0% to 100 % air saturation|
The oxygen sensor spot should be integrated into a dry, clean vessel (humidity or oily residues affect the adhesion and may lead to a detaching of the sensor spot from the vessel wall). The adhesion surface area of the vessel wall should be plane or slightly arched but not strongly curved or corrugated. Otherwise, the spot may detach partly from the vessel wall during the curing process of the silicon glue due to the tension of the spot.
The silicon glue should be cured for at least 60 min. before the test vessel is fill.
Apply a small amount of silicon glue (D5-Spot: ca. 4 µL) onto the red side of the spot.
Position the spot on the plane bent end of a spatula.
Attach the spot to the inside wall of the test vessel.
Shift the sensor spot slightly with little pressure onto the surface so that a small ring of silicon glue emerges around its edge.
No air should be enclosed in the silicon layer between spot and vessel wall.
Silicon stains on the black sensor surface should be avoided since they increase the response time of the sensor.
The light from the blue-green LED excites the oxygen sensor to emit fluorescence. If the sensor spot encounters an oxygen molecule, the excess energy is transferred to the oxygen molecule in a non-radiative transfer, decreasing or quenching the fluorescence signal. The degree of quenching correlates to the partial pressure of oxygen in the matrix, which is in dynamic equilibrium with oxygen in the sample. The decay time measurement is internally referenced.
The Loligo® shuttle tanks are modified versions of the classic operant conditioning chambers (also known as the Skinner box) used for experimental analysis of behavior, e.g. to study operant conditioning and classical conditioning in animals.
An operant conditioning chamber permits experimenters to study behavior conditioning (training) by teaching a subject animal to perform certain actions (like pressing a lever) in response to specific stimuli like a light or sound signal. When the subject correctly performs the behavior, a mechanism delivers food or another reward. In some cases, the mechanism delivers a punishment for incorrect or missing responses.
With this apparatus, experimenters perform studies in conditioning and training through reward/punishment mechanisms. Operant chambers have at least one operandum, that can automatically detect the occurrence of a behavioral response or action.
Typical operanda for primates and rats are response levers. Despite such a simple configuration (e.g. one operandum and one feeder), it is possible to investigate many psychological phenomena in this way.
For this reason, operant conditioning chambers have become common in a variety of research disciplines including behavioral pharmacology, and Skinner's Box have been used extensively for behavioral research in primates and rats.
Loligo® shuttle tanks have been developed for aquatic animals, like zebrafish or crustaceans, and the tank design allows for independent control of water quality in two sub compartments. Tank dimensions are made special to accomodate a wide variaty of animal species and sizes.
Inside the Shuttle tank, the animal can freely "shuttle" between two sub compartments with opposite acting controls.
The computerized Loligo® shuttle systems are equiped with a video camera and lighting conditions enabling real-time pc vision software to detect animal locomotion.
If the animal changes its position from one user-defined zone to the next through locomotion, the computer software (ShuttleSoft) activates/deactivates programmed devices to change environmental conditions inside the tank, e.g. to regulate water temperature to preferred values through behavior. Or you can set up two different (constant) temperature levels in the two tank compartments independent of fish behavior for exposure/avoidance/choice tests.
Today, a main application of Loligo® shuttle tanks measurements of temperature preference in aquatic ectotherms (as well as avoidance behavior), and automated computerized systems, have been made for a range of other environmental factors like water turbidity, salinity, oxygen saturation, pH and pCO2.
The turnkey systems offered include everything needed for video behavior analysis as well as monitoring and regulating water quality.Link
Intermittent flow (or stop-flow) respirometry - Principles and Background
Measurements of oxygen consumption rates on fish and other water breathers commonly involves using one of three different methods:
1. Closed respirometry (or constant volume respirometry)
2. Flow-through respirometry (or open respirometry)
3. Intermittent flow respirometry (or stop-flow)
1. Closed respirometry (or constant volume respirometry)
Measurements in a sealed chamber of known volume (a closed respirometer). The oxygen content of the water is measured initially (t0), then the respirometer is closed and at the end of the experiment (t1) the oxygen content is measured again.
Knowing the body weight of the animal, the respirometer volume and the oxygen content of the water at time t0 and t1, the mass specific oxygen consumption rate (mg O2/kg/hour) can be calculated as follows:
VO2 = ([O2]t0 – [O2]t1) • V/t/BW
An advantage of this method is its simplicity. A disadvantage is that the measurements are never made at a constant oxygen level, due to the continuous use of oxygen by the animal inside the respirometer. This might cause problems when interpreting data, since animal respiration often changes with ambient oxygen partial pressure.
Furthermore, metabolites from the experimental animal, i.e. CO2, accumulate in the water, thus limiting the duration of measurements. This limited time for measurements prevents the experimental animal to recover from initial handling stress that often increase fish respiration significantly and for several hours, thus overestimating oxygen consumption rates.
2. Flow-through respirometry (or open respirometry)
This is a more sophisticated method for oxygen consumption measurements. Experimental animals are placed in a flow-through chamber with known flow rate. Oxygen is measured in the inflow and outflow and oxygen consumption rate (mg O2/kg/hour) can be calculated as:
VO2 = F • ([O2]in – [O2]out)/BW
The advantages of this method are several:
However, this method bring along one significant disadvantage: In order to determine oxygen consumption by open respirometry it is crucial that the system is in steady state. This means that the oxygen content of the in-flowing and out-flowing water, AND the oxygen consumption of the animal, have to be constant.
If the oxygen consumption of the animal for some reason changes during the experiment, steady state will not exist for a while. The above formula will not give the correct oxygen consumption rate until the system is in steady state again. The duration of the time lag depends on the relationship between chamber volume and flow rate. Thus, open respirometry measurements have poor time resolution and are not suitable for determination of oxygen consumption on organisms with a highly variable respiration like fish.
3. Intermittent flow respirometry
Our system for automatic respirometry works by intermittent flow respirometry aiming at combining the best of both 1) closed and 2) flow-through respirometry.
However, the most important advantage is the great time resolution of this method. Oxygen consumption rates of animals can be determined for every 10th minutes over periods of hours or days, making our systems for automatic respirometry extremely suited for uncovering short term variations (minutes) in respiration.
In summary, our system for automatic respirometry is developed for prolonged and automatic measurements of oxygen consumption rate in a controlled laboratory environment.
The automatic measuring proceedure runs in 3 phases:
In the Measuring period (1) the flush pump is off, and the chamber is closed. Fish respiration rate is calculated from the decline in oxygen. During this time the recirculation pump is active to mix the water inside the respirometer and to ensure proper flow past the oxygen sensor.
The measuring period is followed by a Flush period (2) where the flush pump is active pumping water from the ambient temperature bath and into the respirometer. During this period the recirculation pump is inactive and the oxygen curve will raise to approach the level of the amient water.
Finally, the flush pump stops and the loop ends with a short Wait period (3) before starting a new measuring period. This waiting period is necessary to account for a lag in the system response resulting in a non-linear oxygen curve. During the Wait period the recirculation pump is active.
Examples of raw MO2 data
Standard metabolic rate of juvenile Rainbow Trout was determined in static respirometer chambers and with an automated respirometry system from Loligo® Systems. Initial high oxygen consumption rates due to handling stress, were followed by a gradual decline to lower and more stable values indicating standard metabolic rate for the specimen. Notice the high temporal resolution (10 min) of the system revealing sudden changes in MO2.
Courtesy by J.Svendsen/DIFRES & J.Lund/Univ. AarhusLink
It is important that the volume and dimensions of a respirometer chamber fits the body dimensions and respiration rate of the experimental animal.
Temperature affects metabolic rate in ectotherms (like fish). High experimental temperatures thus favor the use of larger chambers and visa versa.
Metabolic rate (or respiration) also scales with the body size of the animals, e.g. large animals have lower mass specific O2 consumption rates than smaller animals (e.g. mouse-and-elephant example).
If the chamber volume is too large, the resulting oxygen depletion curve will be too flat for reliable estimates of the slope that is used in the calculation of the oxygen consumption rate (MO2). A chamber much larger than the animal will allow space for unwanted activity if trying to measure standard metabolic rate of an in-active (static or resting) animal.
On the other hand, a large fish in a small chamber means that oxygen will decline rapidly to hypoxic levels which might affect the metabolism of oxyregulatory species.
As a rule of thumb, we recommend a chamber volume some 10-50 times the volume (≈ wet weight) of the animal depending on the temperature, the species and the size of the specimen, e.g. a 5 L chamber for a large adult 500 g fish at low temperatures (ratio 1:10) or a 500 ml chamber for a small 10 g juvenile fish at higher temperatures (ratio 1:50).Link
Active fish swimming in a tunnel respirometer have high mass-specific oxygen consumption rates (MO2), allowing reliable oxygen consumption estimates in relative higher respirometer volume than for static respirometry. Tunnel respirometers are difficult to build with a fish volume to respirometer volume of less than 1:100. We recommend a maximum ratio between the wet weigth (or volume) of the fish and the volume of the swim respirometer of 1:200, e.g. a 90 liter swim respirometer fits fish down to app. ½ kg. If the volume is too large, the resulting oxygen curve will be too flat for reliable estimates of the slope that is used in the calculation of oxygen consumption rate (MO2).
Another restraint is the dimensions of the test section, which should allow the experimental fish to perform unrestricted swimming. This will largely depend on the species and mode of swimming.
Please contact us to get free advice on which swim tunnel that will be best for your project.Link
To wash out a tube-shaped respirometer chamber requires a volume ~5 times that of the chamber.
Thus, if you use a pump delivering a flow per minute equal to the chamber volume, then a 5 minutes flush period will ensure that 99 % of the water is replaced during each flushing cycle.
Stronger pumps might create too strong a flow forcing fish to struggle or unwanted activity. Lower flows require longer time for flushing, and this will decrease time resolution of MO2 data.Link
Some materials, like Plexiglass/Perspex can act as oxygen stores and sinks, forming pools of oxygen which can be released or stored in a reversible way - see Stevens, J. (1992) J. Appl. Physiol. 72, 801-804.
This can have an important effect when chambers are down-scaled for micro respirometric measurements of very low oxygen fluxes.
Thus, for oxygen measurements in small volumes less than a few millilitres (mL), we recommend chambers of glas and inert components with non or low oxygen storage capacity!
For respirometers with larger volumes (>½ litre), the use of acrylic materials have no significant effects on measurements.Link
If you purchased a software upgrade, please follow the steps below to add new license codes to your WIBU dongle:
If the Input [phase], Oxygen [%air sat] and Temperature [OC] shows NaN (Not a Number), you should check the following:
The NaN should now be replaced by data values.Link
If the Input [phase] and Temperature [OC] shows NaN (Not a Number), you should check the following:
The NaN should now be replaced by data values.Link
If the Board number for the DAQ-BT disappears after having selected it, there could be two reasons:
If the ERROR: Instrument 1 (or 2/3/4) appears, you should check:
Close the error-window.
If the No COM port selected error message appears, you should check that you have selected the assigned COM port for the instrument indicated in the error window; here the first oxygen instrument, a Witrox 1.Link
If the No input for: Ambient oxygen No 1 error message appears, you should check:
If the No input for: Ambient temperature No 1 error message appears, you should check that the Pt1000 temperature probe is connected correctly to the Witrox instrument (red mark on cable must match red mark on Temp input):
If the Input [RPM], Velocity [cm/s] or Output [V] shows NaN (Not a Number), you should check the following:
You should now see data (RPM) values instead of NaN:
The LoligoBT is a 4-fold power strip that is powered from a wall outlet and communicates with a Windows computer via Bluetooth 4.0. LoligoBT is used for wireless software control of equipment connected to one of the four independent electrical sockets, i.e., relays turning pumps, valves, stirrers etc. ON or OFF from a distance.
The Witrox instruments are used for measuring dissolved oxygen using fiber optic mini sensors (optodes) and for measuring temperature.
The DAQ-BT device is used for wireless data acquisition from and control of Loligo® swim tunnels. The built-in Bluetooth communication allows you to place the PC at a distance from the swim tunnel. The device has inputs and outputs for controlling and acquiring RPM data from the swim tunnel motor controller (VFC).
WARNING: DO NOT reset the device to factory defaults!
Resetting the device to factory defaults will void the warranty and remove the pre-installed Loligo® firmware leaving the device unable to communicate with any Loligo® software.
If you have reset the device:
A reset LoligoBT/NETIO4All device must be returned to us for re-installation of Loligo® firmware. Shipping costs will be charged on your account. Please contact our technical support before shipping anything back.
Ways to reset the device (DON’T!!!)
The LoligoBT/NETIO4All device can be reset to factory defaults in two ways:
Contact our technical support if you experience connectivity or performance issue with the LoligoBT/NETIO4ALL. If necessary, we can reset the device for you.Link
The horizontal mini chambers can be fixed in place in an acrylic holder for easier measurements.