Quantification of growth factors in allogenic bone grafts extracted with three different methods
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Quantification of growth factors in allogenic bone grafts extracted with three different methods B. Wildemann Æ A. Kadow-Romacker Æ A. Pruss Æ N. P. Haas Æ G. Schmidmaier
Received: 14 March 2006 / Accepted: 22 May 2006 / Published online: 25 October 2006 Ó Springer Science+Business Media B.V. 2006
Abstract Background Bony allografts are used for defect filling. A reliable sterilization method is the peracetic acid–ethanol sterilization procedure (PES). Several studies showed the antimicrobiological efficacy of this method. Aim of this study was the quantification of growth factors necessary for bone formation in PES sterilized allografts (n = 9). Methods To extract the growth factors from the tissue three different methods were used: (a) use of collagenase 1 for extraction, (b) incubation of the material in a proteinase inhibitor cocktail (Complete), and (c) extraction with guanidine HCl. The supernatants from the different methods were analyzed for the total protein concentration and different growth factors. Results The extraction with guanidine HCl resulted in the highest amount of protein measurable
B. Wildemann (&) Æ A. Kadow-Romacker Æ N. P. Haas Æ G. Schmidmaier Center for Musculoskeletal Surgery, Charite´Universita¨tsmedizin Berlin, Augustenburger Platz 1, D-13353 Berlin, Germany e-mail: [email protected] A. Pruss Institute for Transfusion Medicine (Tissue Bank), Charite´-Universita¨tsmedizin Berlin, Berlin, Germany
in the supernatants of the samples. For comparison of the individual growth factor values the results were normalized to the protein content. The highest growth factor amount/protein was detectable for BMP-2 using the GndHCL method followed by FGFa, IGF-I, TGF-b1, VEGF, and PDGF. Comparing the three extraction methods, significant differences were measured for the individual growth factor content. Conclusion PES sterilized bony allografts contain several growth factors. Depending on the extraction method, the quantity of the analyzed growth factors varies. Keywords Allograft Æ Growth factors Æ Extraction method Æ Collagenase Æ Guanidine HCl Æ Proteinase inhibitor Æ Peracetic acid–ethanol sterilization (PES) Abbreviations BMP Bone morphogenetic growth factor DBM Demineralized bone matrix ELISA Enzyme linked immunosorbant assay FGF Fibroblast growth factor GndHCL Guanidine HCl IGF-I Insulin like growth factor-I PBS Phosphate buffered saline PES Peracetic acid–ethanol sterilization PDGF Platelet derived growth factor TGF-b1 Transforming growth factor-beta1 VEGF Vascular endothelial growth factor
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Introduction Bone defects are a major problem in orthopaedic surgery. The gold standard for defect filling is the use of autogenic bony material. Due to complications associated with the harvesting of autogenic material (Sasso et al. 2005; Silber et al. 2003; Skaggs et al. 2000), allogenic or alloplastic materials might be an alternative grafting material. In the ideal case, this material is biocompatible, not infective, resorbable, osteoconductive, and osteoinductive. Osteoconduction
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