The Effect of Formalin Preservation Time and Temperature on the Material Properties of Bovine Femoral Cortical Bone Tissue

  • Guanjun Zhang
  • Shujing Wang
  • Songyang Xu
  • Fengjiao Guan
  • Zhonghao Bai
  • Haojie MaoEmail author


Literature has reported controversial findings on whether formalin affected bone properties, or not, especially when different preservation time durations and temperatures were involved. Hence, accurately and systematically quantifying the effect of formalin on the mechanical properties of bone using a large dataset is crucial for assessing biomechanical responses based on fixed specimens. A total of 154 longitudinal and 149 transverse cuboid-shaped (12 mm × 2 mm × 0.5 mm) specimens from the midsection of 12 bovine femora from six bovines were prepared and assigned to ten groups, including fresh-frozen, formalin-preserved at 25 °C for 4 weeks and 8 weeks, and formalin-preserved at 4 °C for 4 weeks and 8 weeks. All specimens underwent quasi-static three-point bending tests with a loading rate of 0.02 mm/s. The Young’s modulus, yield stress, yield strain, tangent modulus, effective plastic strain, ultimate stress, and toughness were calculated by optimizing the material parameters to make the force–displacement curve of the finite element prediction consistent with the experimental curve, combined with specimen-specific finite element models. Preservation time and temperature both had significant effects on the Young’s modulus, yield stress, effective plastic strain, yield strain and ultimate stress of cortical bone (p < 0.05). The Young’s modulus, yield stress, and ultimate stress of longitudinal specimens decreased significantly with the increase of preservation time, and the yield strain increased significantly. As the preservation temperature increases, the Young’s modulus of the transverse sample decreased significantly, and the yield strain increased significantly. The preservation time mainly affects the longitudinal specimens, while the preservation temperature mainly affects the transverse specimens. Formalin preservation of bovine femoral cortical bones at a lower temperature and less than 4 weeks is recommended for biomechanical testing.


Cortical bone Formalin fixation Mechanical properties Finite element method Optimization 



This research was funded by the National Natural Science Foundation of China (51205118 and 11402296) and the State Key Laboratory of Advanced Design and Manufacturing for Vehicle Body, Hunan University (51475002). We also acknowledge support from the Canada Research Chairs program.

Conflict of interest

There are no conflicts of interest.


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Copyright information

© Biomedical Engineering Society 2019

Authors and Affiliations

  • Guanjun Zhang
    • 1
  • Shujing Wang
    • 1
  • Songyang Xu
    • 1
  • Fengjiao Guan
    • 2
  • Zhonghao Bai
    • 1
  • Haojie Mao
    • 3
    Email author
  1. 1.State Key Laboratory of Advanced Design and Manufacturing for Vehicle BodyHunan UniversityChangshaChina
  2. 2.Science and Technology on Integrated Logistics Support LaboratoryNational University of Defense TechnologyChangshaChina
  3. 3.Department of Mechanical and Materials Engineering, Faculty of Engineering, School of Biomedical EngineeringWestern UniversityLondonCanada

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