Preparation of L-PRF bone-block & liquid fibrinogen
Introduction
The optimal conditions for periodontal regeneration include: space provision, wound stability, angiogenesis, and healing by primary intention (Susin et al. 2015, Cortellini & Tonetti 2025). An L-PRF bone-block, an upgrade of the so-called "sticky bone", fulfills these requirements. It combines pieces of chopped L-PRF membranes and liquid fibrinogen (both prepared chairside from the patient's blood) with a particulate bone substitute. This combination results in the bone substitute being embedded in a fibrin matrix, formed by the conversion of fibrinogen into a 3D fibrin network, providing a stable and slow-resorbing scaffold. The chopped L-PRF membrane pieces create extra space between the graft particles, facilitating cell ingrowth and neovascularization. Additionally, the L-PRF membranes and liquid fibrinogen contain activated platelets that secrete a wide range of bioactive molecules and growth factors, which play a crucial role in angiogenesis and bone regeneration. Covering the graft with L-PRF membranes further facilitates soft tissue healing over the graft.
SEM figures: showing 3D fibrin network entrapping both the bone substitute and pieces of chopped L-PRF membrane (Castro et al. 2019).
Protocol: step by step
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collect blood for preparation of L-PRF membranes and liquid fibrinogen; the preparation of liquid fibrinogen is performed during the same centrifugation cycle as the L-PRF membranes but requires only 3 min. of centrifugation,
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remove the liquid fibrinogen tube (with an inert inner surface) after 3 min. of centrifugation and replace it with a tube filled with water to balance the centrifuge,
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continue centrifugation of remaining blood tubes for the formation of L-PRF membranes, which require a total centrifugation time of 12 min.,
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aspirate the liquid fibrinogen (yellow upper part in the tube) without red blood cells, using an inert syringe.
STEP 1: liquid fibrinogen:
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collect L-PRF clots and gently compress them to form L-PRF membranes,
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take 2 L-PRF membranes and chop them into very small pieces (2-3 mm).
STEP 2: L-PRF membranes:
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mix the 2 chopped L-PRF membranes with the bone substitute in a small dish,
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thoroughly mix to ensure a homogenous mixture,
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the preferred ratio of bone substitute to chopped L-PRF membranes is approximately 50/50% by volume; typically this equates to 0.5 g of bone substitute for 2 L-PRF membranes,
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for larger defects, you may use 1 g of bone substitute and 4 L-PRF membranes.
STEP 3: Mixture:
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spray the liquid fibrinogen abundantly over the mixture and stir gently for approximately 15 seconds to bring the liquid fibrinogen in contact with the entire mixture,
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the fibrinogen will immediately start to clot into a 3D fibrin network, trapping the biomaterials to form the L-PRF bone-block.
STEP 4: Fibrin network:
Even though the liquid fibrinogen is in a plastic syringe, it will begin to coagulate spontaneously after about 30 minutes (timing is thus critical).
The L-PRF bone-block can be shaped into the desired form during the first minute of mixing, after which it will hold its shape.
Biological concept
The chopped pieces of L-PRF contain a high concentration of activated platelets. These platelets facilitate the conversion of small fibrinogen chains into fibrin polymers by converting prothrombin into thrombin, which subsequently regulates the transformation of fibrinogen into fibrin. The fibrin polymers then form a biochemical matrix with tri-molecular or equilateral junctions, creating a fine and flexible three-dimensional fibrin network. This network entraps the bone substitute and the chopped pieces of L-PRF, resulting in a relatively strong graft with physical characteristics similar to "gummy bears."
step-by-step flow chart
In summary
Video: Preparation L-PRF clot / membrane / plug / exudate
The sequence: 1. gather 2 L-PRF membranes and 0.5 g of bone substitute, 2. chop the L-PRF membranes into very small pieces, 3. mix the chopped L-PRF membranes with the bone substitute to create a homogenous mixture, 4. add the liquid fibrinogen, ensure it penetrates well into the mixture; after one minute, the mixture will transform into a very stable block.
1. Blood collection: Collect 6 blood tubes for the preparation of L-PRF membranes and 2 blood tubes, with an "inert" inner surface, for liquid fibrinogen. The latter requires only 3 min of centrifugation, after which the yellow liquid layer in the tube (up to the red blood cell layer) should be immediately aspirated. Continue centrifugation for the remaining tubes to prepare the L-PRF membranes, which require a total of 12 minutes of spinning.
2. Chop membranes: Chop 2 L-PRF membranes into tiny pieces (± 2-3 mm).
3. Prepare mixture: Combine the chopped membranes and bone substitute in a small dish at a ratio of 2 membranes to 0.5 g biomaterial (corresponding to a volume of approximately 50/50%), ensuring a homogenous mixture.
4. Fibrin network: Spray the liquid fibrinogen abundantly over the mixture and stir gently for approximately 15 seconds to bring the liquid fibrinogen in contact with the entire mixture. The fibrinogen will immediately start to clot into a 3-D fibrin network, which will trap the biomaterials to form the L-PRF bone-block. The mixture can be shaped into the desired form within approximately 1 minute, after which it will hold its shape.
Attention: the liquid fibrinogen will spontaneously coagulate after about 30 minutes; timing is crucial.
When only a small amount of bone graft is required, 1 tube for liquid fibrinogen might be enough; in that case, 7 tubes can be used to prepare L-PRF membranes, and when the liquid fibrinogen tube is removed after 3 minutes, it is replaced by a tube filled with water to keep the centrifuge in balance.
Characteristics of L-PRF bone-block
1: Micro-CT.
A MicroCT image of an L-PRF bone block reveals its composition. Volumetric analyses indicate that the L-PRF bone block consists of 36% bone substitute and 64% L-PRF, which includes pieces of L-PRF membrane and liquid fibrinogen (Castro et al., 2019).
2: Mechanical strength
A well-prepared L-PRF bone-block exhibits high form stability and high elasticity, as demonstrated in the video. Therefore, it is suitable for use in "less contained" defects, such as an extraction socket without a buccal wall or as a graft following a 2-stage sinus floor elevation.
The same concept can be applied to prepare a robust soft-tissue graft by mixing chopped L-PRF with liquid fibrinogen, resulting in an L-PRF soft-block.
paper in preparation
3: Release in growth factors
This figure illustrates the concentrations of several growth factors: Transforming Growth Factor ß-1 (TGF ß-1), Platelet-Derived Growth Factor-AB (PDGF-AB), Vascular Endothelial Growth Factor (VEGF), and Bone Morphogenetic Protein-1 (BMP-1) in L-PRF exudate and liquid fibrinogen.
It also depicts the release of these factors over a 14-day period from an L-PRF bone-block or an L-PRF membrane.
The left column presents the data per time interval (for the liquids immediately after preparation), while the right column displays the cumulative amount over time.
The data show that both the exudate (which comes from squeezing the clots into membranes) and liquid fibrinogen contain these growth factors.
Additionally, it also demonstrates that both the membrane and the L-PRF bone block exhibit a slow release of these growth factors, extending up to 14 days, with the highest release occurring during the first 7 days. This behavior contrasts with PRP, which releases most factors within the first 4 hours.
These concentrations are, however, much lower than the dosage used in clinical/animal trials when applying recombinant human BMP-2, for example.
Key factors for success:
A: Choice of blood tubes
For the preparation of liquid fibrinogen, tubes with an inert inner surface are needed to delay the coagulation process. Despite these measures, coagulation cannot be entirely prevented and typically begins after 30-45 minutes (see right syringe; picture taken after 45 minutes).
This spontaneous coagulation can be delayed by:
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immediate aspiration of the liquid fibrinogen in an inert syringe,
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reducing the temperature by placing the syringe in the refrigerator before use.
B: Timing
Due to the spontaneous coagulation of liquid fibrinogen, it is advisable to follow this time schedule:
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begin with the collection of blood, taking the blood tube for the liquid fibrinogen last,
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remove the liquid fibrinogen tube from the centrifuge after 3 minutes and replace it with a similar tube filled with water,
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continue centrifugation of the remaining tubes for an additional 8-9 minutes,
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collect the liquid fibrinogen as soon as possible,
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collect the other tubes and prepare the L-PRF membranes,
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prepare the L-PRF bone-block promptly and cover the small dish in which it has been prepared to prevent dehydration,
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now you have ample time to perform the surgery at ease.
Important notice
What is the difference between "sticky bone" and an "L-PRF bone-block"?
The preparation of "sticky bone" (introduced by Sohn in 2010 at a symposium in Tokyo) is quite similar to the L-PRF bone-block, with the exception that chopped L-PRF membranes are not added to the mixture. The L-PRF bone-block was introduced by the Department of Periodontology at the Catholic University of Leuven in 2018 (Cortellini et al. 2018)
Short- and long-term resorption after guided bone regeneration
Several studies have tracked, over time, the dimensional changes of the graft following guided bone regeneration (GBR) and reported a significant volumetric loss. This loss occurs in the first months after the augmentation (during the integration/regeneration process), termed "early" resorption, and subsequently in the following years, referred to as "late" resorption (Quirynen et al. 2023). This resorption is attributed to various factors, including the defect morphology, biological principles, and the technique and biomaterials employed.
Recently, Quirynen and co-workers (2023) observed that the individual phenotypic dimensions of the bony envelope predicted the maximal extent of buccal bone regeneration achievable with GBR. This finding also raises questions about the benefits/limits of over-augmentation.
Their comparison of early and late resorption in lateral bone augmentations between an L-PRF bone-block and the gold standard (a composite bone block comprising 50% Bio-Oss and 50% autogenous bone) revealed similar dimensional changes.
What is the difference between liquid fibrinogen and i-PRF?
Injectable PRF (i-PRF) is a flowable formulation prepared without additives (without anticoagulants), using plastic hydrophobic tubes to slow down spontaneous clotting. Blood is centrifuged at 60 g for 3 minutes only, and the upper liquid layer represents the i-PRF.
Liquid fibrinogen is also a flowable formulation again obtained without additives. It is prepared using the same centrifuge setting as for the L-PRF membranes (thus within the same centrifugation procedure, eliminating the need for a second centrifugation/blood collection). Also for liquid fibrinogen plastic hydrophobic tubes have to be used to delay the coagulation cascade, and the spinning time is again 3 minutes with a relative centrifugal force of 340 g (in the area where liquid fibrinogen is created).
Both liquids can be utilized alone or combined with various biomaterials. They can be injected into damaged joints or used for dermatological indications (see Miron & Davies 2025). Compared to the patient‘s whole blood, these liquids contain slightly higher concentrations of platelets and leukocytes.
It is possible to prepare a 100% autogenous bone-block: the L-PRF "dentine" block!
d 0: extraction of a wisdom tooth and removal of all foreign material.
d 0: after grinding the wisdom tooth, both enamel and dentine.
d 0: the application of an L-PRF dentine-block in an extraction socket of an upper first molar of the same patient.
d 0: the graft is covered with L-PRF membranes only.
d 5: suture removal.
m 1: nice healing, but still a minor concavity can be observed.
d 0: RX of the extracted tooth with a loss of the buccal bone plate and an apical pathology.
d 0: these membranes are inserted in the small envelope between the periosteum and the underlying bone.
d 5: the connective tissue is already creeping over the graft.
m 3: image at the day of re-entry.
d 0: the sutures secure the stability of membranes and the graft, no need for primary closure.
d 8: the graft is nearly covered with soft tissues.
m 3: re-entry showed a new buccal bone and an optimal crest dimensions.
m 3: perfect regeneration of the alveolar bone without clear contrast between recipient and regenerated bone.
m 3: hematoxylin & eosin staining of a biopsy taken after 3 months of graft healing, showing bone formation around the dentine particles with intimate contact between both.
Andrade et al. (2020) monitored the healing of L-PRF dentine-blocks in extraction sockets, taking biopsies at 4, 5, and 6 months, respectively. The mean proportions of bone, dentine, and connective tissue were as follows: at 4 months: 26.3, 10.4, and 63.3%; at 5 months: 56.5, 4.8, and 38.7%; and after 6 months: 66.5, 0.9, 32.6%, respectively. Despite the use of enamel and dentine as substitutes, they almost completely disappeared after only 6 months!
It is strongly recommended to clean the tooth mechanically before grinding, removing all amalgam, composite, decay, and calculus, and to disinfect the grind enamel and dentine.
Endodontically treated teeth may contain toxic elements that cannot be eliminated by cleansers and should therefore not be used.
Interesting references
Several videos and/or cases on this webpage are discussed more in detail in the following book: Quirynen M & Pinto N 2022. Leukocyte- and Platelet-Rich Fibrin in Oral Regenerative Procedures. Quintessence Publishing;
ISBN: 978-1-78698-105-9