Inhalation Tower and Controller

Transcription

1 Inhalation Tower and Controller OVERVIEW This manual highlights how DSI s Inhalation Tower and Controller interact as a system. This document will provide an overview of the system components, proper assembly, and basic operation. Data Sciences International TH STREET NW SUITE 100 ST. PAUL, MN USA (651) (800) PN: REV01

2 Table of Contents Welcome... 4 Assumptions... 4 Items currently not covered in manual... 4 Inhalation Tower... 5 Scientific Background... 5 Application Overview... 5 System Components... 6 Hardware Setup... 9 Connecting filter box and drierite tubes... 9 How to set the Inhalation Tower site plugs to the half-open position Using support rods Important safety message Connecting the 14-PORT inhalation tower to the All-In-One controller Installing the air in flow tubing Installing the air out flow tubing Installing the tower Balance Tubing Installing the tower pressure manifold AND TOWER transducer Installing the nebulizer head Installing additional sensors using the multi-purpose sampling manifold Installing the optional temperature/humidity probe Installing the optional photometer Powering up the all-in-one controller Viewing regulation values Viewing temperature and humidity of in flow air Viewing in flow, out flow, and tower pressure Setting regulation values Setting In Flow rate Tower pressure menu Return menu Set aerosol menu P age

3 Aerosol volume menu Aerosol duration menu Calibrate aerosol menu Nebulization Nebulizer Head Starting an experiment Delivering air flow Loading the animal (and acclimation) Maintenance Care and Cleaning Cleaning the Inhalation Tower Cleaning the Restraints/Chamber Cleaning the Nebulizer Head Troubleshooting Technical Support DSI Technical Support North America DSI Technical Support Europe DSI Technical Support All Other Countries Derived Parameters Index P age

4 WELCOME Congratulations on joining the community of users worldwide who rely on DSI s products to perform preclinical physiologic research. Thank you for your interest in DSI products. We are committed to providing you with quality products and services. This manual will help you get to know your inhalation system for rodents. The structure of the manual was designed to sequentially guide you through setting up your DSI system. WHAT YOU WILL BE LEARNING 1. Placement of tower controller and tower in your laboratory. 2. Initial set up of inhalation tower and site plugs (safety notices detailed in manual). 3. Proper assembly of tower controller and inhalation tower components. 4. Turning on the tower controller. 5. Understanding of LCD display and various settings. 6. Nebulization features. 7. Animal handling suggestions. ASSUMPTIONS Prior to use, animals should be properly acclimated to the Allay restrainers. As with any restraint technology, proper acclimation is critical to ensure test subjects are comfortable with restraint and breathing naturally with minimal stress. Please refer to Loading an animal section for further animal handling information. ITEMS CURRENTLY NOT COVERED IN MANUAL The inhalation tower (and tower controller) can be configured in a variety of ways. As such, detail of each configuration is difficult to lay out in the use of a manual. This document will provide an overview of the system components, proper assembly, and basic operation. This manual will not cover the following topics: Integration with external aerosol delivery systems Integration with cascade impactors Integration with smoke generators Use of tower controller app (not always required) Use of plethysmography for measurement of respiratory endpoints If you have questions about any of the above mentioned applications, please contact your DSI sales representative. 4 P age

5 INHALATION TOWER SCIENTIFIC BACKGROUND Since the pulmonary system is considered a vital organ, it is important to have instruments allowing researchers to study the effects of environmental toxins and chemical entities when inhalation is the primary route of exposure. Inhalation research often presents the challenge of greater variability in drug delivery as compared to other administration methods such as intravenous injection or an oral dose. Inhalation Dosing Towers ensure that simultaneous exposure of compounds or gases to the lungs of rodents is well controlled, consistent, and defined. It is crucial that researchers calculate an accurate measurement of the inhaled drug, which is derived by knowing the concentration of the drug delivered as well as the animal s minute volume. Most systems use an assumed minute volume value based on animal species and mass; DSI s inhalation tower provides the ability to integrate plethysmography, allowing for direct minute volume measurement, eliminating the need to make assumptions. Inhalation towers are used in a variety of research disciplines. Inhalation toxicology - study pharmaceuticals, chemicals, and animal health Biodefense Aerobiology - Infections disease, vaccines, and anti-terrorism research Pharmaceutical and contract research organizations assess the impact of a compound on major organ systems or evaluate the safety of new drug candidates Discovery and Academic focus on studies for the proper treatment of respiratory diseases Environmental Toxicology evaluate respiratory health hazard of airborne particulates APPLICATION OVERVIEW The 14-port inhalation tower uses a flow-past design in which there is an inner and outer column. The aerosol or gas is introduced into the inner column and drawn out from the outer column. The subjects are typically arranged as spokes around the tower where they receive air to breathe from the inner column, while the outer column draws away the excess. Each subject has its own port and care is taken to ensure that each port is identical in every way. Symmetry of design is critical. Controlling the exposure is the job of the All-In-One Tower Controller. The tower controller uses two mass flow sensors to monitor and regulate two flows. One flow provides air into the tower, and the other flow draws air out of the tower. The unit regulates the pressure in the outer column to achieve a target pressure which is either slightly above or below zero, as selected by the user. This pressure is monitored in real time, providing instant feedback and uses rapid response valves to allow the excess pressure to vent as needed Patented Allay restraints are used to secure and position each subject identically to help minimize inter- and intra-animal placement variability. Unlike traditional plunger restraints that compromise the test subject s thorax 5 P age

6 and abdomen, the Allay properly secures the subject to create a uniform seal at the exposure port without the need to compress the subject and affect respiratory performance. SYSTEM COMPONENTS Depending on what you purchased, your system may have some or all of the following components: All-in-one Tower Controller Tower pressure manifold Tower pressure transducer Driers and Filters 14-port Inhalation Tower (aluminum or stainless steel) Aerosol block with nebulizer head Support pins for securing heavier animals Patented Allay animal restrainers and optional plethysmographs 4 Flow transducers for plethysmography One calibrator column for plethysmography Optional photometer(s) One optional temperature and humidity sensor The following image is of the front and top panel of the inhalation tower controller: 6 P age

7 The following are the labeled parts of the front and top panel: 1 - The console. This console allows the user to set the target flow rate and outer column pressure. In addition, the user can set the aerosol settings through the console. While the system is running, the console can be set to monitor the flow rate and column pressure, or temperature and humidity. 2 - The plethysmograph calibration port. Plethysmographs are attached to the port with their transducer attached to calibrate the flow signal. 3 - Photometer inputs. These inputs are 3.5mm phone jacks designed to accept analog signals put out by the Casella CEL712 Microdust monitor. 4 - The plethysmograph flow transducer inputs. 4 inputs are provided to monitor ventilation in 4 subjects. 5 - Tower attachments for In flow and Out flow. Inflow is the flow into the tower s inner column. Outflow is the flow of air drawn out of the outer column. 6 - Outer Column Pressure jack. This RJ-45 connector accepts the pressure transducer used to monitor the pressure in the outer column of the inhalation tower. 7 - The Tower Balance connection. A tube should be connected between this tubing port and the outer column of the tower so that the system can regulate the pressure in the outer column. 8 - Nebulizer port. An Aerogen Aeroneb nebulizer can be attached to the port in order to generate aerosol. 9 - Power switch. This is the main power switch to turn the unit on and off Filter Box connection. Part of the Out Flow line to help protect internal components. 7 P age

8 11 - The inflow dryer. This tubing jack allows the user to connect a Drierite tube in line with the in flow to provide dry humidity controlled air The Out Flow dryer. This tubing jack allows the user to connect a Drierite tube in line with the out flow to further protect internal components from moisture. The following image is of the rear panel of the inhalation tower controller: he following are the labeled ports on the rear panel: 1 - USB Connection. This port allows the unit to be connected to a host through USB. Through the USB, the regulating flows, pressures, temperatures, humidity s, and concentrations can be logged at regular intervals. Also, calibrated flow signals can be retrieved in real-time in order to monitor ventilation. 2 - Inlet and outlet ports These ports are required by the pumps and the overflow valve. 3 - The Power Jack. This jack receives a plug from a 12V external power brick. 8 P age

9 HARDWARE SETUP Once you have removed all the components from its packaging, place the All-In-One Controller and the 14-port inhalation tower in a suitable location in your laboratory. The controller weighs approximately 35lbs (15.8kg), while the 14-port inhalation tower (stainless steel and aluminum) weigh approximately 45lbs (20.4kg) and 23lbs (10.4kg) respectively. As a result, the controller and inhalation tower must be placed on a horizontal and sturdy laboratory bench, or similar. Other factors to consider when positioning the system are as follows: Position the inhalation tower, so you will have easy access to all 14 ports, and allow ample surrounding space to connect the Allay animal restrainers. Place the inhalation tower controller under a hood, or make other appropriate accommodations to evacuate the outflow air from the controller, if the compound the test subjects will be inhaling should not be inhaled by laboratory or other personnel. Position the controller so that the exhaust ports (on the back of the unit) are not obstructed. CONNECTING FILTER BOX AND DRIERITE TUBES Drierite tubes are used to dry air going into the inner column (to the animals) and dry air coming from the outer column (from the animals). The filter box is used to further protect internal components in the inhalation tower controller. 9 P age

10 Filter Box Connections: 1) Set the filter box on the top of the inhalation tower controller. 2) Insert the tubing provided into the ports on the side of the filter box. Push until the tubing is firmly seated in the port. 3) Insert the other end of the tubing provided into the filter box connections located on the top lid of the inhalation tower controller. Push until the tubing is firmly seated in the port. Drierite Tube Connections: 1) Locate the two Drierite tubes in the inhalation tower accessory bag. 2) Remove the rubber stoppers. 3) Insert the Drierite tubes onto their respective color coded ports; located on the top lid of the inhalation tower controller. Orientation is not necessary. Caution: Out flow Drierite tubes ARE required (color coded red) to help protect internal components of the inhalation tower controller. 10 P age

11 If in flow Drierite tubes are not desired or used (color coded blue), a bypass tube can be connected. Firmly place the bypass tube into the in flow connection ports on the top of the inhalation tower. HOW TO SET THE INHALATION TOWER SITE PLUGS TO THE HALF-OPEN POSITION It is good laboratory practice before doing any work on the inhalation tower to set all site plugs to the half-open position (described below). The reasons for doing so now, and whenever you are either setting up or shutting down an experiment using the inhalation tower, are as follows: If all site plugs are the closed position, higher in flow rates could cause one or more site plugs to blow out, causing injury to personnel and/or damaging equipment. When loading test subjects into the inhalation tower air should be flowing through the tower. If ports are closed, excess in flow air is being delivered to the test subject. This may prevent the test subject from breathing properly and may result in death. In summary, here are the important take-home messages about the site plugs: Before working on the inhalation tower, always set the site plugs to the half-open position. When your experiment is complete, always return the site plugs to the half-open position. Below is a schematic of the inner and outer column of the inhalation tower. 11 P age

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13 The diagrams below show a site plug in the closed and half-open positions. Closed No air is flowing from the inner column to the outer column. Half-Open Air is flowing from the inner column to the outer column. To half-open a site plug, do the following. 1) Grab the chamber adaptor and pull to remove it from the inhalation tower. 2) Grab the site plug and pull to separate it from the chamber adaptor. 3) Slowly re-insert the site plug into the chamber adaptor until you feel it come to rest in the intermediate, halfopen position. 4) Holding only the chamber adaptor, re-insert the entire configuration into the inhalation tower. 13 P age

14 USING SUPPORT RODS Support rods are required when working with animals larger than mice. Locate the support rods and carefully connect them using the threaded hole above each inhalation tower port. Chambers requiring support rods will have an additional port on the face of the nose piece and can be secured with a thumb screw. NOTE: Ensure the entire system is set up correctly and air flow is being distributed to the tower prior to placing animal in restraints and connecting them to the tower ports. IMPORTANT SAFETY MESSAGE CAUTION. When the inhalation tower is not in use, always set the site plugs to the half-open position. CONNECTING THE 14-PORT INHALATION TOWER TO THE ALL-IN-ONE CONTROLLER This section tells you how to attach the following components to the inhalation tower and controller. In Flow tubing Out Flow tubing Tower balance tubing Tower pressure manifold Tower pressure transducer Nebulizer head and cable Optional sensors such as the temperature/humidity sensor, or the photometer that monitor conditions in the measured and surrounding exposure ports. 14 P age

15 INSTALLING THE AIR IN FLOW TUBING This tubing delivers filtered, dried air to the inner column of the inhalation tower, and evenly distributes it to all 14 ports, where it can be inhaled by the test subjects. For more information, see the Theory of Operation section of the manual. Here s how to install the In Flow tubing: 1) Locate the Y-shaped in flow tubing, which has blue ends and is labeled In Flow. 2) Locate the two In Flow connections on either side of the inner column. These ports have a blue release ring, as shown below. 3) Insert one arm of the Y-shaped tubing into one of the In Flow ports. Push until the tubing is firmly seated in the port. Insert the other arm on the other side. (To remove the tubing from the port, push in the blue ring and pull on the tubing.) 15 P age

16 4) Insert the other (single) end of the tubing into the port on the controller labeled In Flow. INSTALLING THE AIR OUT FLOW TUBING Here s how to install the out flow tubing: 1) Locate the Y-shaped air-out flow tubing, which has red ends and is labeled Out Flow. 2) Locate the two out flow connections on the top of the inhalation tower. These ports have a red release ring, as shown below. 16 P age

17 3) Insert one arm of the Y-shaped tubing into one of the out flow ports. Push until the tubing is firmly seated in the port. Insert the other arm on the other side.. (To remove the tubing from the port, push in the red ring and pull on the tubing.) 4) Insert the other (single) end of the tubing into the port on the controller labeled Out Flow. 17 P age

18 INSTALLING THE TOWER BALANCE TUBING Here s how to install the tower balance tubing: 1) Locate the tower balance tubing, which has a green end and is labeled Tower Balance. 2) Insert the tower balance tubing into the green colored port on the top of the tower. Push until the tubing is firmly seated in the port. 3) Insert the other end of the tubing into the port on the controller labeled Tower Balance. 18 P age

19 INSTALLING THE TOWER PRESSURE MANIFOLD AND TOWER TRANSDUCER The pressure manifold and tower transducer monitor the pressure within the outer column of the inhalation tower. The controller uses these readings to regulate the pressure in the outer column (via the tower balance tubing), and keeps the pressure of the outer column within very close tolerances. Regulating in flow and out flow rates and tower pressure, allow the test subject to breathe as naturally as possible. For more information, see the Theory of Operation section of the manual. Here s how to install the tower pressure manifold: 1) Locate the tower pressure manifold and tower transducer. Pressure manifold 19 P age

20 Tower transducer 2) Insert the pressure manifold into the black-colored sampling port on the top of the tower. Please note that the access hole in the sampling port is fairly small and is not in the center of the port. So, look into the port, and carefully thread the plastic tube into the small access hole. 3) Push the metal part of the pressure manifold firmly into the port, making sure the two black holes face outward and are not obstructed (as shown below) by the any other connections. 20 P age

21 4) Attach the tower transducer to the pressure manifold making sure the transducer cable is pointed up. INSTALLING THE NEBULIZER HEAD The nebulizer head produces a uniform, fine droplet, low-velocity, highly respirable aerosol that is delivered to the test subjects via the inner column of the inhalation tower. For more information, see the Theory of Operation section of the manual. Here s how to install the nebulizer head: 1) Push the nebulizer head into the 22mm hole at the top of the inhalation tower, making sure the connector port is not obstructed. 2) Locate the guide (shown below) on the nebulizer connector, and firmly press the cable into the nebulizer s connector port. (The cable will only fit in the correct orientation.) 21 P age

22 3) Plug the other end of the nebulizer cable into the jack on the tower controller labeled Nebulizer. INSTALLING ADDITIONAL SENSORS USING THE MULTI-PURPOSE SAMPLING MANIFOLD Optional and additional sensors are available. These include a temperature/humidity probe and a 3 rd party photometer. The purpose of these sensors is to provide additional information about the uniformity from one port to another. These sensors measure the following parameters at the surrounding exposure ports. Temperature and humidity of the exposure air. The concentration of a compound delivered to the test subjects. The photometer is capable of detecting the presence of aerosols with approximate concentrations as low as mg/m 3 and as high as 2,500 mg/m 3. The following sections tell you how to install these optional sensors. 22 P age

23 INSTALLING THE OPTIONAL TEMPERATURE/HUMIDITY PROBE The optional temperature/humidity sensor looks like this and uses the multi-purpose sampling manifold. Temperature and Humidity Probe Multi-Purpose Sampling Manifold To attach the temperature/humidity probe, do the following: 1) Remove a standard site plug from one of the chamber adaptors in the inhalation tower. 2) Insert the multi-purpose sampling port into the open chamber adaptor on the inhalation tower. 3) Insert the temperature/humidity probe into the multi-purpose sampling port. 23 P age

24 4) Attach the temperature/humidity probe cable to the jack on tower controller labeled Temperature/Humidity INSTALLING THE OPTIONAL PHOTOMETER The optional photometer attachment for measuring concentration at the exposure sites looks like this. 24 P age

25 To assemble the photometer and the adaptor, do the following. 1) Insert photometer wand into Adaptor A 2) Line up photometer wand so the holes line up with the adaptor 3) Carefully, insert the combined Adaptor B/Manifold through the photometer holes. 4) Insert the blue tubing green and orange colored ports as pictured below. Push until the tubing is firmly seated in the port. 25 P age

26 5) Remove a standard site plug from one of the chamber adaptors in the inhalation tower. 6) Insert the entire assembly into the open chamber adaptor on the inhalation tower. 7) Attach the photometer cable to the jack on the tower controller labeled Photometer Inputs. POWERING UP THE ALL-IN-ONE CONTROLLER Once the appropriate connections have been made between the inhalation tower and the controller, you are ready to power up the controller. The procedure for powering up the controller on a day-to-day basis is exactly the same as the start-up procedure described below. IMPORTANT. Prior to powering up the controller, remove one site plug from the inhalation tower to equilibrate the tower pressure to atmosphere. Pressure may exist if the unit has been recently used. 26 P age

27 Here s how to power up the controller. 1) Ensure one site plug has been removed from the inhalation tower to allow for proper diagnostic checks performed immediately after the unit is turned on. 2) Ensure power supply and power cord are connected to the tower controller and plugged into a power outlet. 3) Locate the main power switch in the bottom-right corner of the controller. 4) Press the top of the switch to turn the power on. The controller s display will light up (as shown below), and will begin running a series of diagnostic tests. 27 P age

28 VIEWING REGULATION VALUES The following sections tell you how to step through the controller menu, so you can view/set the following regulation values. View the temperature and humidity of the In Flow air. View the temperature and humidity of the In Flow air at an exposure port. Set In Flow rate (L/Min). Set the tower pressure (cmh 2 O). Set aerosol volume and duration. Note: To advance through various menus, press Display Select button. VIEWING TEMPERATURE AND HUMIDITY OF IN FLOW AIR In Flow: Displays current temperature and humidity values from air prior to entering the inhalation tower. Tower (if desired): Displays temperature and humidity values from a probe/sampling manifold placed into one of the 14 available ports on the inhalation tower. VIEWING IN FLOW, OUT FLOW, AND TOWER PRESSURE In (Flow): Displays current In Flow rate (L/Min) of air delivered to the inner column of the inhalation tower. Out (Flow): Displays current Out Flow rate (L/Min) of air pulled from the outer column of the inhalation tower. Note: Out Flow rate is non-selectable and is controlled by the selectable In Flow rate. It is automatically set to be no greater than ± 0.25 L/min of the In Flow rate. Pres (Pressure): Displays current Pressure (cmh 2 O ) in the outer column of the inhalation tower. 28 P age

29 SETTING REGULATION VALUES Allows user to enter desired inhalation tower settings for In Flow and Tower Pressure. 1) Press Display Select to get to the Set Regulation menu. 2) Press arrow button to proceed. SETTING IN FLOW RATE Selectable In Flow rate of air delivered to the inner column of the inhalation tower. 1) Press arrow button to adjust. Please note that setting the appropriate air in flow rate depends on the following factors: The species you are testing. 29 P age

30 The number of test subjects connected to the inhalation tower. The number of ports used for sampling equipment. RECOMMENDED VALUES FOR START-UP AIR FLOW The recommended values for start-up air flow depend on how many exposure ports are being used (animals + sensors), as discussed in the example above. Please select the appropriate start-up air flow that suits your particular experimental situation from the table below. Please note that these are starting points to consider. Your particular application may require/demand a different flow setting. Care must be given to provide sufficient air flow to the animals without causing injury or death. Once you have determined the approximate total flow required, adjust the in flow rate on the tower controller. Due to incremental settings on the in flow rate menu, the calculated flow required may be approximate. Total Number of Exposure Ports Being Used (Animals + Sensors) Mouse Rat All flow rates are expressed in liters per minute (L/min) Mouse Approximate start-up airflow = 0.3 L/min x n Rat Approximate start-up airflow = 0.5 L/min x n Where n = total number of exposure ports being used (animals + sensors) Example In this example, two of the exposure ports in the inhalation tower are equipped with a temperature/humidity probe and a photometer. In addition, eight rats will be used in this example. 30 P age

31 The total number of exposure ports required for our experiment is 10 (eight rats + two sensors), meaning we need a total In Flow rate (as shown by the table below) of 5.0 liters per minute into the inner column. Note: The 4 additional exposure ports will not be used. As required by this experimental set-up, it is important to place the 4 unused site plugs into the closed position. Closed No air is flowing from the inner column to the outer column. TOWER PRESSURE MENU The Tower Pressure menu allows users to enter the desired pressure setting in the outer column of the inhalation tower. There are three typical settings: a) Slightly negative b) Slightly positive c) No pressure Selectable in increments of 0.25 cmh 2 O with a range of ±1.00 cmh 2 O. 1) Press Display Select to get to Set Regulation menu. 2) Press arrow button to proceed. Press arrow button to adjust. 31 P age

32 Note: The most common setting is a slightly negative pressure (-0.5 cmh 2 0) RETURN MENU To return to the previous menu, press arrow button. SET AEROSOL MENU The Set Aerosol menu allows users to enter the volume of aerosol to be delivered and its duration. 1) Press Display Select to get to Set Aerosol. 2) Press arrow button to proceed. AEROSOL VOLUME MENU The Set Aerosol Volume menu allows users to enter the volume of aerosol that is to be delivered via the inner column of the inhalation tower. Incremental settings by 0.1 ml. 32 P age

33 1) Press arrow button to adjust. AEROSOL DURATION MENU The Set Duration menu allows users to enter the desired duration of the aerosol being delivered. Incremental durations by 1 minute. 1) Press arrow button to adjust. CALIBRATE AEROSOL MENU Aerogen nebulizer heads provide the best means for introducing controlled compound aerosols into the inhalation tower. Over time the Aerogen nebulizer head s ability to produce aerosol degrades. Proper care can extend the life, but will eventually the performance will degrade to the point that replacement is necessary. As the performance degrades, the system needs to correct for this degraded performance by applying more power to get the same aerosol. This is achieved by recalibrating the nebulizer head. The tower controller saves the latest calibration information. Press Display Select to get to Calibrate Aerosol menu. 1) Press arrow button to proceed 33 P age

34 2) Load ml of liquid, by pipette or syringe, into the nebulizer head. Hold the nebulizer head in your hand. Do not block the bottom. 3) Press arrow button to begin 4) Watch the liquid nebulize. When all the liquid is gone (when mist stops coming out of the nebulizer head) press arrow button to stop the timer. 5) A new, accurate rate will be displayed. 6) Press the arrow button to accept this rate, or the Display Select button to reject this rate and test again. The nebulizer is now calibrated and the rate is saved. For the gold neb heads, the rate should be close to 0.3 ml/min. For the purple neb heads the rate should be close to 0.1 ml/min. 34 P age

35 NEBULIZATION NEBULIZER HEAD The nebulizer head may be autoclaved and should be cleaned between each use. It consists of a cup, a piezoelectric element, and an aperture plate with holes. When energy is applied, the plate vibrates rapidly, creating a micro-pumping action that generates a fine particle aerosol. It is capable of aerosolizing a broad range of formulations, including solutions, suspensions, small molecules, and macromolecules. We offer two different versions of nebulizer heads, one purple and one gold, pictured here : 2.5 to 4µm (purple) has a smaller particle size and a rate of approximately 100 µl/min : 4 to 6µm (gold) has a larger particle size and a rate of approximately 300 µl/min. For precise nebulizer head efficiency rate determination, please refer to Calibrate Aerosol Menu section. STARTING AN EXPERIMENT After you have configured the system, you are ready to start an experiment. Before loading any animals into restrainers and placing them on the inhalation tower ports, always ensure the site plugs are in the half open position. DELIVERING AIR FLOW Start delivering air to the inhalation tower by pressing the In Flow button on the tower controller. A green light on both the In Flow and Out Flow button will appear. Air will begin to flow through the inhalation tower and the tower controller will start to monitor the system. It may take a few moments to properly balance the system. 35 P age

36 LOADING THE ANIMAL (AND ACCLIMATION) This section depicts the various steps required to properly position and load the animal into an Allay Restraint chamber. Practice and confidence will allow a single investigator to easily perform this procedure. Note: Animal position in the chamber may vary based on species and strain of animal being used. Acclimation practices may vary. However, DSI recommends increasing acclimation time on an incremental basis. In addition, the steps outlined below may be conducted in phases, allowing the animal to become more comfortable with the restraint technique. CONSIDERATIONS Prior to any animal restraint, place the restraint tube in the animal cage; allowing the animal to explore on its own. The investigator should start with the largest restraint collar and work toward an appropriate size. Once comfortable with the process and the typical size of the animals used, choosing the appropriate collar will become easier O-rings can be lubricated using silicone grease to make it easier to insert/remove components. Place a small dab on your fingertip and run your finger around the entire O-ring. Do not leave a lot of greasy residue on the O-ring. A little grease goes a long way If animals will be connected to an apparatus, it is suggested that hardware and software systems (if applicable) are ready for operation prior to placing the animal in the Allay restraint chamber SUGGESTED ANIMAL HANDLING PROCEDURE 1. Pick up the animal by the base of the tail. 2. Load the animal into the rear of the Allay restraint and attempt to walk it along toward the open end 36 P age

37 3. Once it reaches the other end of the restraint, apply more tension to the tail to hold the animal in place and ensure it does not travel too far. Use your judgment to decide when the animal is in a good position to drop the collar in place a. Allow the animal to become acclimated to this position 4. Holding the animal and restraint in one hand, use your other hand to pick up one of the restraint collars 5. Carefully place the restraint collar over the animal s neck, behind the ears, and in front of the shoulder blades. Push the collar all the way down to secure the animal. Be careful not to catch any fur or skin when pushing down the collar a. If the animal is able to back out, the collar is too big. Proceed to the next size collar NOTE: Animals will inherently resist the process. Always observe the animal s behavior for any signs of significant stress and respond accordingly. 6. Once the appropriate sized collar has been properly placed, use an index finger to keep the Allay restraint collar in place. a. Allow the animal to become acclimated to this restraint/position 37 P age

38 7. Next, locate the nose piece that contains the latex seal (may differ based on species). Place it in your dominant hand place, ensuring the non-concentric hole is facing down Position of mouse (rats will not have eyes protruding beyond latex seal) 8. Slowly move the nose piece toward the animal. Allow the animal to adjust and find its way out through the hole, watching closely not to catch the animal s feet or whiskers a. A fresh latex seal is preferred so it will be more compliant b. After prolonged use, latex seals will harden and will need to be replaced 9. A properly placed nose cap is verified by ensuring the latex seal fits snugly over the nose and mouth and beyond the whiskers a. Allow the animal to become acclimated to this restraint 38 P age

39 10. For mice, place the inhalation nose piece on the end of the restrainer and over the collar. When placing the restraint on the inhalation tower, care should be taken to ensure the restraint is properly attached and the animal remains in a safe position. For additional information on acclimation, please contact DSI and request/refer to the Allay Restraint Instructions document. MAINTENANCE Replace the tubing as needed, when it becomes visibly clogged, or cloudy with aerosol deposition. Tubing should be free from cracks, moisture, and deposits. The inhalation tower controller is outfitted with a filter box and is considered a consumable item. The frequency of replacement is dependent on the amount of use/exposure of the system. Therefore, it is difficult to provide an exact timeframe. It is recommended that an extra filter box be on hand in case it needs to be exchanged. The filter box (and filters) help protect sensors inside the inhalation tower controller from any drug/moisture that is coming back into the system through the Out Flow tubing. They main indicator as to when a filter box should be replaced is when the inhalation tower controller has difficulty regulating/controlling the Out Flow rate. If it is having difficulty, a red light indicator will appear near the Out Flow button on the front of the inhalation tower controller. 39 P age

40 The Drierite tubes are filled with desiccant to remove moisture from the air flow. The Drierite crystals used in the cylinders are originally blue in color. When the crystals turn pink, they must be replaced. Crystals may be replaced in three different ways: 1) install a new cylinder of Drierite crystals available from DSI; 2) remove and replace the crystals with new Drierite crystals; or 3) reuse the existing crystals. If using plethysmographs, replace pneumotach screens in the plethysmograph as needed. A starter set of replacement screens is included in your chamber maintenance kit. Replacement kits for inhalation tower controller consumables are available In Flow Dryer Kit Out Flow Dryer Kit Filter Box Kit Accessory Kit Tubing Kit (included in accessory kit) Pressure Manifold Kit (included in accessory kit) CARE AND CLEANING CLEANING THE INHALATION TOWER The inhalation tower can be disassembled for proper cleaning. Using the top and bottom latches, the top plate, inner core, outer core, and base can be separated from one another. The top plate can be removed by releasing the three latches near the top of the inhalation tower. 40 P age

41 The outer core can be removed from the base by releasing the three latches near the bottom of the inhalation tower. Carefully lift the outer core from the base to reveal the inner core of the inhalation tower. To remove the inner core, carefully lift it from the bottom plate. The inner core (newer versions) has an adaptor plug located on the underside. This plug prevents the black base plate from coming in contact to exposure elements. If desired, the adaptor plug can also be removed. To remove, locate the threaded plug tool in the accessory kit. Carefully, thread the tool on to the adaptor plug. Once secure, simply pull the adaptor plug out. 41 P age

42 If desired, the bottom plate of the inhalation tower can be removed from the black base by removing 4 screws located on the underside. 42 P age

43 Depending on the type of exposure, end users may wish to autoclave components. The following is a list of components that can be autoclaved: Top Plate Inner Core Outer Core Bottom Plate Site Plugs Chamber Adaptor DO NOT autoclave the (black) base of the inhalation tower. CLEANING THE RESTRAINTS/CHAMBER Clean animal chamber of debris as needed by wiping with a cloth or rinsing with warm water. For a more thorough cleaning, use warm water and a mild dish detergent. It is best to let the chambers thoroughly air dry. If required, dry with a cloth towel. DO NOT submerge the plethysmograph portion of the chamber (if purchased) as this will affect pneumotach screens and the reference chamber. DO NOT USE WINDEX or window cleaner to clean the plethysmograph. Continued use of Windex will eventually crack the lucite material. DO NOT put the chamber in the dishwasher. CLEANING THE NEBULIZER HEAD Clean the nebulizing equipment after each use. To clean the nebulizer head, remove it from the plethysmograph or aerosol block and rinse it in warm, soapy water. Let it soak for a few minutes. It can be submerged. Do NOT use a high pressure water stream; let it run gently into the device. Do NOT touch the metal membrane or attempt to brush it, as this may cause damage, but do rinse it from both the top and the bottom Do NOT clean the head with alcohol. Alcohol will damage the metal membrane. It is best to use soapy water for all general cleaning. 43 P age

44 CORRECT DRYING POSITION To dry, shake off the excess water and wipe gently with a paper towel or Kimwipes. Set the head on its SIDE in a dry place. INCORRECT DRYING POSITION Do NOT stand the head up or place it in any inline circuit while not in use. Trapped humidity may cause the metal membrane to discolor. If you have a stubborn protein build up on the head, you may try Efferdent. Dissolve a tablet in a glass of water. Then filter the resulting solution with a simple syringe filter. Nebulize it for 10 minutes. Rinse thoroughly and air dry. You may pat gently with paper towels. DO NOT leave the head attached to the distribution reservoir overnight after use. If you have been using a pathogen and wish to decontaminate, simply disconnect the head for the power cord, then autoclave it, using the flash steam sterilization cycle, 134C for 4 minutes. CAUTIONS To avoid mechanical or electrical damage, do not drop the nebulizer head or the control module. Do not use in the presence of devices generating high electromagnetic fields such as magnetic resonance imaging (MRI) equipment. Disconnect the nebulizer head from control module before cleaning. Do not immerse or autoclave the nebulizer control unit. Use only with components specified by Aerogen. Inspect all parts before use, and do not use if any parts are missing, cracked or damaged. In case of missing parts, malfunction or damage, contact your DSI representative. Do not use or store outside of specified environmental conditions. 44 P age

45 TROUBLESHOOTING TECHNICAL SUPPORT Getting technical support from DSI will, we hope, be a quick and painless process, because we are strongly committed to helping you our users get the very best from our products. In other words, when you hit a road block, our aim is get your experiment, monitoring program, or whatever back up and running as quickly as possible. We are here to help! DSI TECHNICAL SUPPORT NORTH AMERICA Support@datasci.com Toll-free in U.S. and Canada Phone: Monday through Friday: 8 AM to 5 PM CST (except Holidays) DSI TECHNICAL SUPPORT EUROPE Europe-support@datasci.com Phone: Monday through Friday: 8 AM to 5 PM CET DSI TECHNICAL SUPPORT ALL OTHER COUNTRIES Phone: P age

46 DERIVED PARAMETERS The following parameters are reported from the Inhalation Tower Controller: Parameter Units Description Conc in mg/m 3 Concentration from the photometer placed near the inner column Conc port mg/m 3 Concentration from the photometer placed at a port P cmh 2 O Pressure in the outer column T in C Temperature measured as the flow exits the Inhalation Tower Controller H in % Humidity measured as the flow exits the Inhalation Tower Controller T port C Temperature measured at the port H port % Humidity measure at the port F in L/min Mass flow rate of air entering the tower (exiting the Inhalation Tower Controller) F out L/min Mass flow rate of air exiting the tower (not counting the overflow) P baro mbar Current barometric pressure Using FinePointe software and optional plethysmographs, common pulmonary parameters can be derived from a respiratory flow signal: Parameter Term Units Description f Frequency BPM The instantaneous, breath-by-breath rate of breathing TV Tidal Volume ml or L The inspired volume of air per breath. The integral of the negative section of the flow curve MV Minute Volume ml/min or L/min The product of the tidal volume and the respiratory rate, calculated on a breath-by-breath basis AV Accumulated Volume ml or L Total volume breathed over a specific period of time. Calculated by adding the TV for each breath to the previous Accumulated Volume. Used in Dosimetry experiments, AV provides a good measure of the actual dose an animal receives The maximum inspiratory flow that occurs in one breath PIF Peak Inspiratory Flow ml/sec or L/sec PEF Peak Expiratory ml/sec The maximum expiratory flow that occurs in one breath Flow or L/sec Ti Inspiratory Time Sec The time spent inhaling during each breath, from the start of inspiration to the end of inspiration (determined by interpolation of start of expiration). The time flow is negative Te Expiratory Time Sec The time spent exhaling during each breath, from start of expiration to end of expiration (determined by interpolation back to zero). The time flow is positive dv Delta Volume ml or L The difference between inspiratory and expiratory volume EF50 The flow at the point 50% of TV is ml/sec or L/min An indicator of bronchoconstriction 46 P age

47 expired Tr Relaxation Time Sec The time it takes for the animal to expire a certain amount of volume Rinx Rejection Index % Calculates the percentage of breaths rejected 47 P age

48 INDEX A Animal Acclimation 36 Application Overview 5 C Calibrate Aerosol 33 Care and Cleaning 40 Connections 14 D Derived Parameters 46 F Flow Rates 30 H Hardware Setup 9 L Loading the animal 36 M Maintenance 39 N Nebulization 35 Nebulizer Head 35 O Optional Sensors 22 P Power 26 R Return Menu 32 S Scientific Background 5 Set Aerosol P age

49 Setting Regulation Values 29 Starting an experiment 35 System Components 6 T Tower Pressure Menu 31 Troubleshooting 45 V Viewing Regulation Values P age