Platelet-rich fibrin (PRF) therapy has been widely applied in regenerative dentistry, and PRF preparation has been optimized to efficiently form fibrin clots using plain glass tubes. Currently, a shortage of commercially available glass tubes has forced PRF users to utilize silica-coated plastic tubes. However, most plastic tubes are approved by regulatory authorities only for diagnostic use and remain to be approved for PRF therapy. To clarify this issue, we quantified silica microparticles incorporated into the PRF matrix. Blood samples were collected into three different brands of silica-containing plastic tubes and were immediately centrifuged following the protocol for advanced-PRF (A-PRF). Advanced-PRF-like matrices were examined using a scanning electron microscope (SEM), and silica microparticles were quantified using a spectrophotometer. Each brand used silica microparticles of specific size and appearance. Regardless of tube brands and individual donors, significant, but not accidental, levels of silica microparticles were found to be incorporated into the A-PRF-like matrix, which will be consequently incorporated into the implantation sites. Presently, from the increasing data for cytotoxicity of amorphous silica, we cannot exclude the possibility that such A-PRF-like matrices negatively influence tissue regeneration through induction of inflammation. Further investigation should be performed to clarify such potential risks.

Because of its simple operation, platelet-rich fibrin (PRF) is becoming more popular than the original form, platelet-rich plasma (PRP), in regenerative dentistry. PRF preparation requires plain glass blood-collection tubes, but not either anticoagulants or coagulation factors. However, such glass tubes designed for laboratory testing are no longer commercially available. Although several glass tubes specifically designed for PRF preparation are available, many clinicians prefer to obtain stably supplied substitutes, such as silica-coated plastic tubes produced by major medical device companies. The quality of PRF prepared by silica-coated tubes has not been assessed and we previously reported significant contamination of silica microparticles in the resulting PRF matrix and alerted clinicians against the use for PRF preparation. To further assess the biosafety of the silica microparticles, we presently examined their effects on human normal periosteal cells derived from alveolar bone. The periosteal cells were obtained from explant cultures of small periosteal tissues obtained from healthy donors. Silica microparticles were obtained from silica-coated tubes and added to cell cultures. Cellular responses were monitored using a tetrazolium assay, phase-contract inverted microscopy, an immunofluorescence method, and scanning electron microscopy. Silica microparticles adsorbed onto the cell surface with seemingly high affinity and induced apoptosis, resulting in significant reduction of cell proliferation and viability. These findings suggest that silica microparticles contained in plastic tubes for the purpose of blood coagulation are hazardous for various cell types around sites where silicacontaminated PRF matrices are implanted.Publisher's Note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Platelet-rich fibrin (PRF) is a fibrin matrix enriched with platelets. The PRF matrix is thought to form a steep gradient of platelet density around the region corresponding to the buffy coat in anticoagulated blood samples. However, this phenomenon has not yet been proven. To visualize platelet distribution in PRF in a non-invasive manner, we utilized near-infrared (NIR) imaging technology. In this study, four types of PRF matrices, bio-PRF, advanced-PRF (A-PRF), leukocyte-rich PRF (L-PRF), and concentrated growth factors (CGF) were compared. Blood samples collected from healthy, non-smoking volunteers were immediately centrifuged using four different protocols in glass tubes. The fixed PRF matrices were sagittally divided into two equal parts, and subjected to modified immunohistochemical examination. After probing with NIR dye-conjugated secondary antibody, the CD41+ platelets were visualized using an NIR imager. In L-PRF and CGF, platelets were distributed mainly on and below the distal surface, while in bio-PRF and A-PRF, platelet distribution was widespread and homogenous. Among three regions of the PRF matrices (upper, middle, and lower), no significant differences were observed. These findings suggest that platelets aggregate on polymerizing fibrin fibers and float up as a PRF matrix into the plasma fraction, amending the current “gradient” theory of platelet distribution.

Aim Platelet‐rich fibrin (PRF) matrices are compared with regard to their ability to retain and release growth factors. Although this ability is thought to influence regenerative outcomes, it remains unclear how it is regulated. To address this question, we compared advanced PRF (A‐PRF) and concentrated growth factor (CGF) matrices in terms of distribution of platelets, transforming growth factor‐β1, platelet‐derived growth factor‐BB, vascular endothelial growth factor and matrix metalloprotease‐9 (MMP9). Methods Blood samples were obtained in glass tubes and immediately centrifuged to prepare A‐PRF or CGF matrix according to their specific protocols. Both matrices were compressed, embedded in paraffin and subjected to immunohistochemical examination. Results Leukocytes and plasma proteins were localized on the proximal surface including the interface corresponding to buffy coat. In A‐PRF, platelets were distributed homogenously, while growth factors and fibronectin were localized on the distal surface and MMP9 was mainly colocalized with leukocytes. In CGF, in contrast, platelets were localized on and below the proximal surface like leukocytes, growth factors were diffused homogenously and MMP9 was found in the plasma protein layers. Conclusion Although these preparations do not allow accurate quantification, platelet counts and growth factor levels seemed higher and leukocytes were less activated in A‐PRF. This may explain A‐PRF’s higher ability to release growth factors.

Platelet-rich fibrin (PRF) matrices were originally prepared using plain glass tubes without the aid of coagulation factors because coagulation factor XII is activated by glass surfaces. Recently, the use of silica-coated plastic tubes as a substitute of glass tubes has been recommended for PRF preparation. This recommendation is owing not only to the shortage of glass tubes for medical use in the market, but also the higher coagulation activity of silica-coated plastic tubes and equal quality of PRF. However, these matrices are not the same. To evaluate the differences, we compared glass-and silica-coated plastic tubes in terms of platelet distribution and quantity in concentrated growth factors (CGF). CGF matrices were immediately prepared from freshly collected blood samples, fixed after red thrombus removal, and divided into two equal pieces sagittally. One piece was used for CD41 detection and the other was applied as an isotype control. Platelet distribution in CGF matrices was examined, without embedding or sectioning, by a novel method using invisible near-infrared imaging. The dehydrated membranous CGF matrix was more transparent. Thus, the fluorescence signal was clearly detectable with less scattering. Platelets were distributed mainly in the distal side of the glass-prepared CGF matrix, but homogeneously in the silica-prepared CGF matrix. Platelet count was positively correlated with fluorescence intensity. Although not yet fully developed, this imaging technique enabled us to recognize the differences in platelet distribution and quantity in CGF matrices by excluding bias caused by the technical limitations of scanning electron microscopy and conventional immunohistochemical methods.