A major objective of biomaterial research and tissue engineering is to promote a material-induced tissue reaction that leads to regeneration and an effective wound-healing process in the defective area. Thus, a biomaterial should serve as a temporary barrier to cover defects and promote tissue regeneration while being tissue compatible and, most importantly, clinically applicable. In the field of tissue regeneration, vascularization plays a crucial role as it ensures a continuous supply of nutrients to and the removal of waste products from the scaffold and the transplanted region.

Concepts such as the use of a biomaterial alone^1,2  or preseeded with different primary mesenchymal³  or endothelial cells^4,5  are usual prerequisites for clinically applicable tissue engineering. However, concepts involving the precultivation of cells require time for cell isolation or cultivation as well as the possibility to aseptically handle more complex constructs in the operating room. These become major challenges if there is demand for a fast, robust, and “simple” approach by means of cell-based tissue engineering. Obviously, time is one of the most precious commodities in the clinic.

To ensure that methods for tissue engineering are widely applicable in the clinical field, it is necessary to modify them in a way that they are readily available and relatively easy to use within the daily clinical routine. Therefore, the steps between the preparation and application have to be minimized and optimized to make practical implementation realistic. Thus, it is the overall goal to develop concepts that are of natural origin, can be produced “close” to the patients, and would expedite the process of implantation while being financially realistic for the patient and the health system.

The requirements mentioned above led us to look for new strategies in which a new class of biomaterials is generated from autologous blood.^6–9  Platelet-rich plasma (PRP) was the first generation scaffold derived from human blood samples, and this concept was widely studied and established with common denominators such as the addition of anticoagulants and bovine serum and double/-centrifugation.¹⁰  Comparative studies showed that PRP has a positive effect on the wound-healing process and tissue regeneration.¹⁰  However, the addition of anticoagulants and bovine serum limits the clinical application of PRP and calls for alternative, clinically feasible strategies.

With the aim of improving and streamlining these preparation methods—especially towards cell-seeded biomaterials generated from the patient's own blood—a concept of platelet-rich fibrin (PRF) was developed.^11–16  This fibrin scaffold, which does not possess any cytotoxic potential,¹⁷  is obtained from 9 mL of the patient's own blood after 1 centrifugation step. The factors previously used in the preparation of blood-based scaffolds, such as anticoagulants or bovine serum, were excluded from this preparation procedure, which minimized the risk of trans-contamination. The main approach was to keep the methodology convenient and applicable for clinical use.^11–16  The three-dimensional fibrin network is capable of mimicking the extracellular matrix in terms of its structure,^18,19  which creates the environment for cells to function optimally.

Choukroun's PRF is derived from human blood and contains a variety of blood cells—including platelets, B- and T-lymphocytes, monocytes, stem cells, and neutrophilic granulocytes—as well as growth factors.^20,21  A major advantage of this method is its simplicity of preparation. The centrifugation process activates the coagulation process and as a result the clot is formed. This clot consists of a 3-dimensional fibrin network in which the platelets and other blood cells are entrapped. The release of growth factors from the PRF clot commences 5 to 10 minutes after clotting and continues for at least 60 to 300 minutes.^21,22  Several studies with Choukroun's PRF have shown the tissue regenerative potential of this cell-loaded 3-dimensional scaffold. Interestingly, this scaffold is also a carrier for mesenchymal cells. Ling He et al were able to show that different cell types, such as rat osteoblasts, could differentiate and proliferate when cultured on the leukocyte-rich PRF (L-PRF). Differentiation and proliferation rates were investigated in terms of transforming-growth factor β (TGF-β), platelet-derived-growth factor AB (PDGF-AB), and alkaline phosphatase (ALP) activity at 5 time points. The study showed a slow but constant release of TGF-β1 and PDGF-AB as well as ALP activity.²³ 

Further experimental studies demonstrated that even dermal pre-keratinocytes, human gingival fibroblasts, pre-adipocytes, and maxillofacial osteoblasts underwent differentiation and proliferated with Choukroun's PRF. Dental pulp cells were also able to grow and undergo further “differentiation” on the fibrin scaffold.²⁴  In addition to the experimental approaches discussed above, clinical applicability of the material was tested. Mazor et al showed that biopsies of the augmented/implant area taken 6 months after implantation revealed new bone formation, thus highlighting its possible osteo-inductive potential.²⁵ 

Although the studies mentioned above underline the considerable potential of PRF in term of tissue regeneration and clinical application, it was still not clear how cells are distributed in this type of scaffold depending on varying centrifugation time and speed (ie, cumulative centrifugational force). The aim of the study was to assess histologically and histomorphometrically the cell distribution pattern of fibrin clots by applying the standard PRF (S-PRF) protocol with 12-minute centrifugation time and 2700 rpm as well as the new advanced PRF (A-PRF) protocol with 14-minute centrifugation time and 1500 rpm.