Proximal Cadaveric Femur Preparation for Fracture Strength Testing and Quantitative CT-based Finite Element Analysis
March 11th, 2017
We present a robust protocol on how to carefully preserve and prepare cadaveric femora for fracture testing and quantitative computed tomography imaging. The method provides precise control over input conditions for the purpose of determining relationships between bone mineral density, fracture strength, and defining finite element model geometry and properties.
The overall goal of this protocol is to facilitate the consistent preparation of human cadaveric femora for computed tomography, or CT scanning and mechanical testing in a sideways fall on the hip configuration. This method can help answer questions in the field of osteoporosis research about bone imaging and bone biomechanical properties. The main advantage of this technique is that it is a comprehensive procedure for preparing proximal femurs in a reproducible manner for bone fracture research.
Begin by covering four or five pound rice bags with plastic bags to prevent their contamination and protect the DEXA surface with plastic lined paper. Transfer two of the plastic wrapped rice bags onto the scanner table. Then lay two 24 hour room temperature thawed femora on top of the rice bags.
Such that the proximal ends, including the femoral heads, are centered on the bags with the posterior sides facing down. Cover the exposed anterior approximal femur end with the other two rice bags, and position the machine head over the approximal femur. Now select the femur exam in the machine software interface, and position the arm on top of the cadaveric femurs.
After analyzing the bone marrow density, an automatic T-Score will be generated classifying the bone as normal, osteopenic or osteoporotic. Before beginning the bone modification sanitize the workspace with 70%isopropyl alcohol, and cover the table with absorbent paper pads with plastic film on one side. Then place the entire femur into a custom made cutting fixture with the head of the femur against the acrylic plate of the fixture.
Holding the diaphysis against the two pins on the cutting fixture, tighten the slotted plate on to the diaphysis. When the bone is secured, pass the cast cutter through the slotted plate as a guide, and incise the distal shaft of the femur. Now remove the bone from the fixture, and use a 25 millimeter deep curette to clean the marrow from the medullary cavity.
As needed use a sponge gauze to clean the interior surface of the bone. Then grip the bone with a dry cloth and drill a 10 millimeter hole through the distal end of the bone, approximately 25 millimeters from the proximal cut end of the specimen. Before beginning the bone potting procedure, label the potting container with the appropriate bone identification details, and adjust the embedding fixture to the proper orientation.
Place the potting container in the base of the embedding fixture inside a fume hood, and place the bone into the potting container. Then align the neck with the pointer in the fixture to adjust the internal rotation angle of the bone to the desired value. Next, mix 60 grams of PMMA powder with 30 grams of liquid resin, in a disposable cup compatible with PMMA until the powder has dissolved.
When the mixture is pourable, decant the PMMA solution into the potting container, until the container is about half full. And allow the PMMA to set for 10 to 15 minutes inside the fume hood. While the mixture is curing, carefully wrap the bone in saline saturated towels to prevent tissue dryness.
Periodically check the femur to ensure the bone remains aligned in the container during the curing process. After the resin has cured, completely fill the potting container with 90 grams of PMMA and liquid resin, and allow the PMMA to cure for another 10 to 15 minutes. When the additional resin has cured, rewrap the bone in saline soaked paper towels, and fully cover the bone with a plastic bag.
Before x-ray imaging, move the beam head to the desired height and place an unexposed cassette on the cart under the beam. Next place the femur on top of the cassette in the medial lateral position, and move behind the lead line portable wall for personal protection. Label the specimen images, and then use the trigger to expose the bones.
To scan the bones by computed tomography, secure a completely thawed plastic bag wrapped femur in the calibration phantom in the CT scanning fixture. Use the system laser guides to confirm that the middle of the fixture is properly aligned with the longitude in the laser of the CT scanner. Moving the table up or down until the horizontal laser rests between the head and trochanter as appropriate.
Then press the laser on/off button to recheck the fixture alignment with the CT lung access laser, and press the zero button to zero the table position. When the bone is in place use standard CT operating procedures to scan the bone with the appropriate scanning parameters, obtaining an image slice thickness of 0.4 millimeters and a pixel size of 0.30 to 0.45 millimeters depending on the size of the field of view. Bone mineral density scanning using a DEXA scanner as just demonstrated, allows measurement of the total hip and neck bone density.
As well as acquisition of the T-Score for each specimen. X-ray images obtained from this method can be used to discern the presence of prior fractures or diseases, such as cancer, that might affect femoral strength. CT images can be used to obtain three-dimensional bone geometry and volumetric bone mineral distribution to be used in quantitative CT-based finite element analysis.
While attempting this procedure it is important to remember to minimize the number of freeze style cycles, and to keep the femur moist during preparation steps. After watching this video, you should have a good understanding of how to prepare a femur for consistent imaging and testing of a sideways fall on the hip configuration.
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Overview
This protocol outlines a method for the preservation and preparation of human cadaveric femora for fracture testing and CT imaging. It aims to enhance the understanding of bone mineral density and fracture strength in osteoporosis research.
Key Study Components
Area of Science
- Bone biomechanics
- Osteoporosis research
- Computed tomography imaging
Background
- Understanding bone properties is crucial for osteoporosis research.
- Fracture testing helps in assessing bone strength.
- CT imaging provides detailed insights into bone structure.
- Standardized preparation methods are essential for reproducibility.
Purpose of Study
- To develop a reliable protocol for preparing cadaveric femora.
- To facilitate consistent imaging and mechanical testing.
- To investigate the relationship between bone density and fracture strength.
Methods Used
- Covering rice bags to prevent contamination.
- Using plastic lined paper to protect scanning surfaces.
- Positioning thawed femora on prepared surfaces for scanning.
- Conducting mechanical tests in a sideways fall configuration.
Main Results
- Establishment of a reproducible preparation method.
- Improved accuracy in measuring bone properties.
- Enhanced understanding of the biomechanical behavior of femora.
- Insights into the impact of osteoporosis on bone strength.
Conclusions
- The protocol provides a comprehensive approach to femur preparation.
- It supports research into osteoporosis and related conditions.
- Future studies can build on this method for further insights.
Frequently Asked Questions
Executive Industry Relevance
This protocol enables reproducible preparation of human cadaveric femora for biomechanical testing and imaging, supporting preclinical evaluation of bone-targeted therapeutics. By preserving tissue integrity and standardizing geometry and mineral density assessment, it enhances predictive confidence in fracture risk models used in osteoporosis drug development. The method facilitates translational continuity from ex vivo testing to in silico modeling via QCT/FEA, aiding mechanistic de-risking of skeletal agents.
Strategic Applications in Biopharma R&D
Early Discovery & Target Validation
- Scientific Value: Enables functional validation of bone strength phenotypes in human tissue, supporting target hypothesis testing for osteoporosis pathways.
- Operational Value: Provides a standardized workflow to minimize biological variability across specimens, improving assay consistency.
Screening & Assay Development
- Scientific Value: Generates quantitative CT-derived bone mineral density and geometry data suitable for assay calibration and biomarker alignment.
- Operational Value: Uses calibrated phantoms and standardized scanning parameters to ensure reproducible imaging outputs across batches.
Translational & Preclinical Research
- Scientific Value: Supports validation of QCT/FEA models by providing ground-truth mechanical test data, enabling predictive de-risking of compound effects on bone integrity.
- Operational Value: Facilitates cross-functional collaboration between imaging, biomechanics, and modeling teams through standardized specimen preparation.
Pipeline & Workflow Integration
The method integrates into the discovery continuum by enabling ex vivo biomechanical assessment that informs target validation and lead optimization in bone metabolism research.
- Discovery Biology: Supports hypothesis testing of genetic or pharmacological modifiers of bone strength through controlled mechanical testing.
- Screening: Delivers reproducible BMD and geometric readouts that enable quantitative comparison of experimental conditions.
- Analytics: Provides high-resolution CT data and failure load measurements for computational model training and validation.
- Translational Research: Bridges ex vivo findings to in silico predictions via QCT/FEA, supporting extrapolation to human fracture risk.
- Enterprise Reuse: Establishes a scalable, standardized preparation platform applicable across multiple bone-targeted programs over extended timelines.
Operational & Enterprise Impact
- Scientific Value: Enhances predictive confidence in bone efficacy models by reducing mechanical testing variability and preserving native tissue properties.
- Operational Value: Ensures reproducibility through standardized fixturing, temperature control, and handling protocols across large specimen cohorts.
- Strategic Value: Improves go/no-go decision quality by providing human-relevant biomechanical data early in the discovery pipeline.
- Portfolio Impact: Enables risk-adjusted prioritization of skeletal therapeutics based on validated fracture strength endpoints.
Implementation Considerations
- Requires expertise in bone biomechanics, CT imaging, and PMMA handling for proper specimen alignment and embedding.
- Dependent on access to calibrated CT scanners, DEXA densitometers, and mechanical testing systems configured for sideways fall simulations.
- Necessitates standardized labeling, thawing cycles, and moisture control to prevent specimen degradation during preparation.
- Must account for donor variability in age, sex, and bone health status when interpreting mechanical outcomes across studies.
- Limited to ex vivo applications; does not replace in vivo pharmacokinetic or pharmacodynamic profiling of bone-active agents.
Biopharma FAQs
Chapters in this video
0:05
Title
0:47
Dual X-ray Absorptiometry (DXA) Bone Mineral Density Measurement
1:53
Distal Bone End Cleaning, Cutting and Drilling
3:13
Bone Potting
5:13
X-ray and Computed Tomography (CT) Bone Imaging
6:43
Results: Representative Bone Imaging and Analyses
7:28
Conclusion
