4pt Bending, Mouse This protocol is for standard Jepsen 4pt bending of adult mouse bone. Safety considerations Please reference the Jepsen laboratory when using this protocol. This protocol is subject to change. We hold no responsibility for errors in the protocol or user error that lead to personal or property damage. Please follow all federal, local, and institutional rules and regulations when using this protocol. Personal Protective Equipment Wear nitrile gloves, lab coat, and safety glasses as a minimum protection, unless otherwise indicated. Potential Hazards Hydraulic testing systems have the potential to do great bodily harm if used improperly. Never have any part of your body below the actuator when the hydraulics are on. Materials Several cotton swabs (to use dowels for practice test) 50lb Sensotec load cell (S/N 997569) External LVDT (050 MHR) Small frame with extra set of springs Jepsen 4pt mouse fixtures Squeeze bottle of 1X Phosphate Buffered Saline (PBS) Glass petri dish Small pair pickups 4x4 gauze Method Equipment Set-Up 1. Set-up small frame with a set of extra springs 2. Mount the load cell and connect to the proper load cell cable 3. Mount the external LVDT and connect to the DTR-451 input cable 4. Make sure the points are aligned properly. See the image below for properly aligned points. a. Set the brass point first. b. While holding one of the lower points, loosen the screw (barely) for that point. If you don t hold the point that you are loosening, it will rotate, which causes alignment issues. Hold the point in position as you tighten it! You want the gap between the two bottom points to be consistent from front to back. c. If you are having issues with positioning, spin the bottom two points around. They are easier to align from one direction. d. Make sure all 4 screws in the image are tight before testing begins. 4pt Bending, Mouse Page 1 of 6 Revision 1
MTS System Set-Up 1. Turn on computer 2. Turn on the Teststar box (Switch is located on the back-top right) 3. Turn on DTR-451 (Digital Transducer Readout) 4. Turn on main hydraulics in pump room (turn to low first, then to high) 5. Log-in to computer (user: ORL user; password: trabeculae) 6. Double Click Station Manager Icon 7. Select a configuration file (ex. External_lvdt.cfg) 8. Select a parameter set (ex. ext_lvdt) 9. Click Open 10. Select MPT icon from left task menu 11. Display Signal Auto Offset by selecting the first button under the Station Controls (this field box contains the input signals for the parameter set you selected and allows you to zero any of the inputs by selecting the button in between the current value and offset boxes) 12. Create a new output folder for the day in the directory C:\MTS_Specimen_Data (every user should create their own folder under their name or the study name and then organize data by date) 13. Display the MPT Specimen Editor by clicking the third button from the left on the MPT control panel (it looks like 3 dumbbells) and set the Specimen Directory Path to the folder for the day that you just created in step 12 (ex. C:\MTS_Specimen_Data\User\StudyName\TodaysDate). 14. Close Specimen Editor 15. Reset hydraulic interlocks on the Station control panel. Select the HSM Low button, then the High button. This can also be done on the Remote Station Controller. 16. Select Enable Manual Contol on the Remote Station Controller and find the center of the actuator s traveling distance (+5 mm). Zero the axial displacement on the Signal Auto Offset panel. 4pt Bending, Mouse Page 2 of 6 Revision 1
17. Move the MTS crosshead to within your working distance and then tighten down all the bolts. 18. Record the PID values of the system. Go into Station Setup, Channels-Axial. The PID values should be in the first tab. System Warm-Up 1. System gets warmed up at the beginning of each test day to get rid of transient changes in temperature, performance, etc. Warm-up consists of running a cyclic program for an hour. 2. Open the warmuplarge Procedure by selecting the open procedure button, which is the first from the left on the MPT control panel, and select the warmuplarge file 3. Create a new specimen called warmup. To create a new specimen, click on the second button from the left on the MPT control panel (it looks like a single dumbbell). No data will be written to this warmup file, but the system needs a specimen to proceed 4. Do not have any fixtures below the actuator during warmup. 5. If Enable Manual Command is selected, deselect it (On the Remote Station Controller). 6. Select the Play button on the MPT Station Control panel and wait an hour. LVDT Calibration 1. While the system is warming up, is the most time efficient to run the LVDT calibration. 2. Select the LVDT in which you want to use (Jepsen lab uses 050 MHR.) 3. Mount the LVDT rod and cylinder into an axial micrometer stage. 4. Disconnect the Teststar cable from the DTR-451 output and connect it to a volmeter. 5. Make sure the LVDT cable is not connected to the DTR-451 and turn the Zero adjustment on the DTR-451 to reach a 0.00 reading on the voltmeter 6. Reconnect the LVDT cable to the DTR-451 and move the LVDT core with the micrometer stage so that the readout is as close to 0.00 as possible on the voltmeter. 7. Zero the micrometer stage display and select the mm increment button to work in metric units. (This is the null position of the LVDT from which the plus and minus displacements are usually measured) 8. Move the LVDT core from the null position to the LVDT plus full scale position in the positive direction (ex. +1mm) using the micrometer stage adjustment. (You should set this range based on your test design.) 9. Turn the Span adjustment to reach a display reading of approximately +10 Volts on the voltmeter. (This should correspond to approximately 10,000 on the DTR-451 display) 10. You may need to adjust the Phase control next to obtain a maximum display reading of +10 Volts, otherwise skip this step. 11. Displace the LVDT core back to the precise null position with the micrometer stage adjustment. If necessary adjust the Zero control until the voltmeter display reads 0.00 Volts. 12. Displace the LVDT core to the negative full scale position (ex. -1mm) using the micrometer stage adjustment. 13. Turn the Span adjustment to reach a display reading of approximately -10 Volts on the voltmeter. (This should correspond to approximately 10,000 on the DTR-451 display) 14. Repeat Steps 8-13 as necessary until the plus and negative full range positions coordinate as closely as possible to the +\- 10 Volt display on the voltmeter. 15. If the directionality of the core appears reverse it is okay as the directionality is based on which direction you have the rod relative to the cylinder. 4pt Bending, Mouse Page 3 of 6 Revision 1
16. Move the micrometer in equal increments over the full range (positive and negative) and record the voltage output on the voltmeter at each increment. Make sure to have at least 10 data points. 17. Use excel to record and graph the data points so that you can calculate the calibration factor in mm/volts (Based on your range you should be able to determine that your calibration factor is close to the ideal calibration factor) For example, if your range was +/-1mm your ideal calibration factor is (2 mm / 20 Volts =.1 mm/volts). 18. Mount the LVDT rod and cylinder into your test set-up. 19. Reconnect DTR-451 output cable to Teststar cable Standard Testing 1. Several small dowels should be tested at the beginning of each day to confirm good data collection and point alignment. 2. Open the SpecialK procedure from the MPT panel. This program ramps down to -3 mm displacement at a rate of 0.05 mm/sec. Data is acquired at a frequency of 25 Hz and is sent to an output file (ex. Specimen.dat) in your directory. Open the Procedure Editor in the MPT panel to confirm these settings haven t changed. 3. Open the Scope window. Ensure that it is displaying load and the external LVDT. 4. Zero the load readout by selecting the Auto Offset button in the Signal Auto Offset window. 5. Move the actuator down (make sure you select Enable Manual Control on the Remote Station Controller) until the upper points overlap the lower points. 6. Create a new specimen in the MPT panel. It is important that you always create a new specimen for each specimen or else data may get combined into one output file. 7. Load the dowel between the points ensuring contact with all 4 points. 8. Slowly move the actuator up/down to get good contact. In rare cases, adjusting the brass point on the top may be necessary. Make sure that the dowel is snug in place, without trying to apply any pre-load. 9. Once the dowel is in position, disable the Manual Control on the Remote Station Controller. 10. Adjust the LVDT height until you are in the proper range. Make sure that you are starting in the correct positive or negative position ensuring that you will stay in range throughout your test. 11. Zero both the axial displacement and external LVDT readings on the Signal Auto Offset window. 12. Select the Play button on the Station Control panel. This begins the data acquisition. 13. A Start button will appear in the bottom of the MPT panel. Pressing this will begin the actuator movement. 14. Repeat for all standards. If anything appears odd in the scope window, make any adjustments as needed. Specimen Testing 1. Open the SpecialK procedure file from the MPT panel 2. Zero the load readout from Signal Auto Offset window. 3. Create a new specimen using the identification numbers of your testing samples. 4. Load your sample a. Place the anterior side of the bone in tension. b. Center the mid-diaphysis of the bone between the upper two points. c. Bring the actuator down slowly so that the bone is in contact with all 4 points. In rare cases, you may need to loosen the brass points at the top. d. The bone should be snug, without having a pre-load on it. 4pt Bending, Mouse Page 4 of 6 Revision 1
e. If having difficulties positioning the sample, remove it, place in PBS, and try again. It is critical to keep the bone hydrated, as a drying bone will have different mechanical properties. f. If there is still a lot of soft tissue on the bone, gently remove it with gauze. Rehydrate before loading the bone into the fixture. 5. Check that the external LVDT is still reading in an appropriate range as the dowels can be different diameters from the bone samples. As the LVDT has a ±1mm range, make sure the DTR- 451 box is reading -9500 at the minimum, so that as loading commences, it will move towards 10000. Place the bone sample back in PBS, and confirm that the LVDT is reading in range. 6. Secure your sample into the test set-up 7. Disable the Manual Control on the Remote Station Controller. 8. Zero both the axial displacement and external LVDT readings on the Signal Auto Offset window. 9. Select the Play button on the Station Control panel. This begins the data acquisition. 10. A Start button will appear in the bottom of the MPT panel. Pressing this will begin the actuator movement. 11. Repeat for all samples 12. Record the position of the break for every sample. If anything outside of the ordinary happens during a test (rotation of a bone sample, slipping, etc) note it next to the test. MPT Procedure Editor Any test can be set-up with the MPT Procedure Editor wizard which is found by selecting the Procedure Editor button on the MPT panel (it is the fourth from the right). The process palette (the button that looks like a painter s palette) associated with this window allows you to drag and drop different types of test commands, data acquisition inputs, events, etc. If you double click on the button associated with the procedure type it will pop up a new window for you to enter the test parameters. Make sure to always include a data limit detector that will interrupt your test command if the limit is reached. Base your limit on the type of load cell you are using. Make sure that if you start with a previously existing procedure and edit it that you save it as a NEW procedure and DO NOT save over the existing procedure unless you are the person who created it. Shutting Down 1. Before turning off the hydraulics, move the actuator up and remove the small testing frame from below the actuator. Then turn off the hydraulics. The actuator drifts after the pressure is turned off, so you want to make sure it doesn t run in to anything. 2. Close the Signal Auto Offset Window and Station Manager. 3. Answer the series of questions as follows: Do you really wish to quit? YES You have changed the station parameters.do you want to save these changes? NO Save the current test parameters? NO 4. Turn off the Teststar box and DTR-451. 5. Log-Off Certification 4pt Bending, Mouse Page 5 of 6 Revision 1
I have read and understand the above SOP. I agree to contact my Supervisor or Lab Manager if I plan to modify this procedure. Name Signature Trainer s signature Date Principal Investigator / Safety Officer Date (signature) Printed name: Designated Trainer Date (signature) Printed name: 4pt Bending, Mouse Page 6 of 6 Revision 1