From: Release kinetics of the model protein FITC-BSA from different polymer-coated bovine bone substitutes
Step 1 | Coarse degreasing of femur bone marrow with a steam jet |
Step 2 | Thorough degreasing of femur bone marrow by application of chloroform:methanol (3:1) solution for three consecutive times at 24 h, 72 h and 24 h intervals. |
Step 3 | Washing with distilled water, bleaching by application of hydrogen peroxide [H2O2] (3% for 15 min), washing with distilled water. |
Step 4 | Demineralization by application of 0.5 M (M) hydrogen chloride [HCL] for three consecutive times at 1.5 h intervals. |
Step 5 | Washing with distilled water, sawing out the cuboid femur slices, application of 0.5 M HCL for 1 h, washing with distilled water, inactivation of osteoinductive matrix proteins by application of 4 M guanidine-HCL/Tris*-HCL (pH value 7.0) for 24 h at 4 °C. |
Step 6 | Application of 50 mM Tris-HCL/150 mM sodium chloride [NaCl] (pH value 7.0), washing with distilled water for 30 min, temporary freezing at − 80 °C for 72 h, lyophilization at − 4 °C and 0.05 mbar for 24 h, storage at 4 °C until gamma sterilization and final sterile storage in 50 ml containers at room temperature (RT). |
Step 7 | Assorting of ICBM with medium cancellous bone density for use in this study. |