Clinical, Radiographic, and Histomorphometric Outcomes Following Ridge Preservation Using Leukocyte Platelet Rich Fibrin (L-PRF) and Freeze-Dried Bone Allograft (FDBA) Layered Technique: A Prospective, Randomized Clinical Trial
Phase: NA
Conditions: Ridge Preservation
Sponsor: University of Alabama at Birmingham

 Leukocyte Platelet Rich Fibrin (L-PRF) and Freeze-dried 
Bone Allograft (FDBA) Layered Technique vs L-PRF/FDBA 
and L-PRF Alone in Influencing Quantity and Quality of 
New Bone Formation in Grafted Extraction Sockets 
 
 
Study Protocol & Statistical Analysis Plan 
 
 
NCT04133363 
 
January 31, 2023 
 
 
Nicolaas C. Geurs, DDS, MS, Principal Investigator 
 
University of Alabama at Birmingham 
Birmingham, Al 35294 
 
 

 
 
1 
 
Clinical, Radiographic, and Histomorphometric Outcomes Following Ridge 
Preservation using Leukocyte Platelet Rich Fibrin (L-PRF) and Freeze-Dried 
Bone Allograft (FDBA) Layered Technique: A Prospective, Randomized 
Clinical Trial 
Protocol 
Ver. 1 
1.0 Investigational Design 
 
1.1.Study Design 
This investigation is a randomized controlled trial of three techniques to evaluate clinically, 
radiographic, and histologically the efficacy of  L-PRF with freeze-dried bone allograft (FDBA) 
layered technique in improving quantity and quality of new vital bone formation and alveolar 
ridge dimensional stability following ridge preservation in comparison to L-PRF alone and L-
PRF/FDBA.  
 
Control technique: Ridge preservation using L-PRF/FDBA and ridge preservation using 
L-PRF alone 
 
Test technique: Ridge preservation using L-PRF/FDBA layered technique 
1.2. Objectives:  
- To evaluate clinical and radiographic dimensional changes at alveolar ridge 3 months after 
ridge preservation procedure using L-PRF alone, L-PRF/FDBA, and L-PRF/FDBA Layered 
technique. 
- To evaluate quantity and quality of new vital bone in sockets 3 months after ridge preservation 
procedure using L-PRF alone, L-PRF/FDBA, and L-PRF/FDBA Layered technique. 
 
1.3. Hypotheses and Specific Aims:  
• Hypothesis 1: Ridge preservation using L-PRF/FDBA Layered technique will best preserve 
alveolar ridge dimensions in comparison to using L-PRF alone.  
• Hypothesis 2: Ridge preservation using L-PRF/FDBA Layered technique will improve new 
vital bone formation at the middle and apical thirds of the socket in comparison to using L-
PRF/FDBA.  
 
• Specific Aims:  
 
1. To compare quantitative clinical alveolar ridge dimensions (in mm) 3 months after ridge 
preservation between the three groups using an intraoral scan and direct clinical assessment. 

 
 
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2. To compare quantitative two- and three- dimensional radiographic alveolar ridge dimensional 
changes (in mm) 3 months after ridge preservation between the three groups using Cone 
Beam Computed Tomography (CBCT) and virtual implant planning software.  
3. To compare qualitative and quantitative histologic characteristics of new bone between the 
three groups after 3 months of ridge preservation by quantifying the distribution (in %) of 
new vital bone, connective tissue, residual graft particles, and artifact at the healed socket 
site. 
2.0. Study Population 
 
2.1 Entrance Criteria 
Patients presented to the postdoctoral periodontal clinic at UAB School of Dentistry with a 
treatment plan for tooth extraction and implant placement will be screened according to the 
criteria. Thirty healthy adult eligible patients (ten in each group) of any ethnicity or gender may 
be enrolled into the study.   
Participants must be at least 18 years old with demonstrated ability to understand and consent to 
the proposed study procedures. Decisionally-impaired adults and/or minors who cannot consent 
for themselves will not be enrolled.  
Smokers (>10 cigarettes/day) patients or women who report a current pregnancy or patients who 
are not systemically healthy enough to receive out-patient elective dental care at the time of 
enrollment will be excluded.  
Participants must have one or more hopeless single-rooted tooth that require extraction and 
planned to be replaced with a dental implant, with healthy adjacent teeth not planned for 
extraction. The extraction socket should have residual 4 bone walls following atraumatic tooth 
extraction. 
Patients must demonstrate high compliance, adequate oral hygiene, and denied history of alcohol 
or drug abuse.  
3.0 Study procedures 
 
3.1 Summary of study procedures 
• Screening/baseline visit: Upon enrollment into the study the patient's medical history and 
electronic records will be reviewed. Patients will be treatment planned by an 
interdisciplinary team consisting of a surgeon and a prosthodontist or restorative dentist. 
Clinical examiners will conduct clinical and radiographic exams to determine eligibility 
according to the above inclusion criteria. An intraoral scan will be taken to perform clinical 
measurements. Study visits and objectives will be explained to all participants. IRB 
approved written informed consent will be obtained from all the participants. 
 
• Visit 1 (Surgical visit): Extraction and ridge preservation 

 
 
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• Randomization: Participants will be stratified and randomized to one of three treatment 
groups: Table 1, by permuted block randomization approach to ensure same number of 
patients in each group, using computer-generated random number list.  
 
 
Table 1. Groups of treatment. 
Group 
Treatment 
Number of 
participants 
Group 1 
 
Atraumatic tooth extraction followed by ridge preservation using 
Leucocyte and Platelet Rich Fibrin (L-PRF) + Freeze-Dried Bone 
Allograft (FDBA) Layered technique 
10 
Group 2 
 
Atraumatic tooth extraction followed by ridge preservation using 
Leucocyte and Platelet Rich Fibrin (L-PRF) + Freeze-Dried Bone 
Allograft (FDBA) 
10 
Group 3 
 
Atraumatic tooth extraction followed by ridge preservation using L-
PRF 
10 
 
• Surgical procedure: A loading dose of prophylactic antibiotic will be dispensed 
(Amoxicillin 2 grs or Clindamycin 600mg, 30 minutes to one hour prior to the surgery). 
Venous blood will be drawn (antecubital fossa) with a 25-butterfly needle to fill 4-8 
collection tubes of 10ml each. The tubes will not contain any additive or anticoagulant. 
One tube will be centrifuged for 3 minutes and the other tubes will be centrifuged for 12 
minutes.  After centrifugation, L-PRF clots will be collected and stored in a closed 
sterile storage container that enables compression of clots into membranes/plugs. 
Additionally, the one tube that received less centrifugation (fibrinogen) will be used as 
an additive with the L-PRF membranes and bone allograft. The bone graft material 
utilized for patients in group 2 and 3 will be MinerOss cortico-cancellous freeze-dried 
bone allograft (FDBA).  
L-PRF Preparation  
 
 
 
Venous blood collection 
Centrifuge white-covered 
tube for 3 minutes and red-
covered tubes for 12 minutes 
White-covered tubes: 
Fibrinogen 
Red-covered tubes: L-PRF 
membranes 

 
 
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After preparation and isolation of the surgical area, local anesthesia will be accomplished by 
infiltration and/or intraoral nerve block. Intravenous moderate conscious sedation or inhalation 
sedation will be used in patients who requested it.  
Tooth or teeth will be gently elevated using a minimally traumatic technique that prevent loss 
of supporting bone. Socket(s) will be curetted, irrigated with saline, and inspected following 
extraction and verified for the presence of intact walls.  
A Cone Beam Computed Tomography (CBCT) scan will be taken immediately after the tooth 
extraction, before ridge preservation procedure. It will serve for baseline radiographic 
measurements. 
Group 1: L-PRF/FDBA Layered technique:  
After socket is cleaned, a L-PRF plug will be formed, placed and compacted at the apical and 
middle third of the socket. Two membranes of L-PRF will be cut into small pieces and 
combined with 0.5 cc of cortico-cancellous bone allograft (FDBA), in a 50%-50% proportion, 
and fibrinogen forming sticky bone. The sticky bone will be placed and compacted at the 
coronal third of the socket (over the L-PRF plug already placed). A membrane of L-PRF will 
be placed at the coronal extent of the socket, flaps will be reapproximated and sutured with 
Monocryl 5-0 sutures. 
Group 1: L-PRF/FDBA Layered technique  
 
 
L--PRF membranes 

 
 
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L-PRF plug formed and placed 
in the apical and middle third 
of the socket 
 
L-PRF membrane chopped 
into small pieces and 
combined with FDBA and 
fibrinogen forming sticky bone 
 
Sticky bone is placed at the 
coronal third of the socket 
(over the L-PRF plug) 

 
 
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A L-PRF membrane is placed 
covering the graft and sutured.  
  
Group 2: L-PRF/FDBA: 
After socket is cleaned, L-PRF membranes will be chopped into small pieces and combined with 
0.5 cc of cortico-cancellous bone allograft (FDBA) and fibrinogen forming sticky bone. The 
sticky bone will be placed and compacted at the along the socket. A membrane of L-PRF will be 
placed at the coronal extent of the socket, flaps will be reapproximated and sutured with 
Monocryl 5-0 sutures. 
Group 3: L-PRF 
After the socket is cleaned, the socket will be completely filled with L-PRF plugs and a L-PRF 
membrane will be placed at the coronal extent of the socket, flaps will be reapproximated and 
sutured with Monocryl 5-0 sutures. 
Post-surgical analgesics will be prescribed and/or dispensed as deemed necessary by the treating 
surgeon and a continued course of antibiotic will be prescribed for a 1-day time of period. 
Written and oral home care instructions will be provided. 
• Visit 2 (Follow-up visit): Sutures will be removed after two weeks. Surgical site will be 
evaluated for healing status and post-operative instructions on resuming oral hygiene 
measures will be given to patients. Medical history will be reviewed, and patients will 
complete a questionnaire regarding their assessment of the post-operative discomfort they 
experienced during the initial healing phase, including number of days of altered 
activity/discomfort and number of tablets of pain medications taken.  
 
• Visit 3 (Follow-up visit): After four weeks, surgical site will be evaluated for healing status 
and oral hygiene instructions will be reinforced. Medical history will be reviewed, and 
adverse events will be record. 
 

 
 
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• Visit 4 (Surgical visit): Bone biopsy and implant placement  
After three months, surgical site will be evaluated. A second intraoral scan will be taken to 
evaluate clinical ridge dimensions. A second CBCT scan will be taken to evaluate the 
dimensional changes of the healed socket and plan the optimal implant location using 
implant planning software. Medical history will be reviewed, and adverse events will be 
record. 
A surgical approach will be performed to take a bone biopsy for research purposes and 
implant placement at the same site. The procedure will be performed under local anesthesia 
and IV conscious sedation if patient requested it. After a full-thickness mucoperiosteal flap 
will be raised, the ridge dimensions will be measured at the crestal bone for dimensional 
changes in height and width using a calibrated surgical caliper and UNC-15 periodontal 
probe. Prior to implant placement, a bone biopsy will be taken from the center of the new 
bone using a 2mm internal diameter trephine bur. The dimensions of the harvested bone core 
will be 2mm in diameter and the length will be according to the implant dimension to be 
placed. The biopsy will be stored in 10% neutral buffered formalin and will be sent to the 
Histomorphometry and Molecular Analysis Core in the UAB Lyons Harrison Building. The 
Implant preparation osteotomy site will be completed following manufacturer protocol.  
 
• Visit 5 (Follow-up visit): Sutures will be removed two weeks after the bone biopsy and 
implant placement. Surgical site will be evaluated for healing status and post-operative 
instructions on resuming oral hygiene measures will be given to patients. Medical history 
will be reviewed and patients will complete a questionnaire regarding their assessment of the 
post-operative discomfort they experience during the initial healing phase, including number 
of days of altered activity/discomfort and number of tablets of pain medications taken.  
 
• Visit 6 (Follow-up visit): After four weeks, surgical site will be evaluated for healing status 
and oral hygiene instructions will be reinforced. Medical history will be reviewed, and 
adverse events will be record. 
 
• Visit 7 (Final study visit): In six months, surgical site will be evaluated for healing status. 
This evaluation will include a clinical examination, periapical radiograph, intraoral scan, and 
assessment of overall oral health and hygiene. Patients will complete a questionnaire 
regarding their satisfaction with study participation. Patients will be offered a dental cleaning 
at no charge at this visit. Medical history will be reviewed, and adverse events will be record. 
 
 
 
 
 

 
 
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Table 2. Study schedule of events 
Procedure Events 
Screening 
Baseline 
V1 
Extraction 
& ridge 
preservation 
V2 – V3 
2-4 wks 
Follow-
up 
V4 
3 months 
Bone biopsy 
and implant 
placement 
V5-V6 
2-4 wks 
Follow-
up 
V7 
6 
months 
 
IRB-Consent 
x 
 
 
 
 
 
Medical history 
review 
x 
x 
x 
x 
x 
x 
Inclusion/exclusion 
criteria 
x 
 
 
 
 
 
Oral exam 
x 
x 
x 
x 
x 
x 
Randomization to 
either group 
 
x 
 
 
 
 
Tooth 
extraction/Ridge 
preservation 
 
x 
 
 
 
 
Bone core biopsy 
 
 
 
x 
 
 
Implant placement 
 
 
 
x 
 
 
X-ray 
x 
 
 
x 
 
x 
CBCT 
 
x 
 
x 
 
 
Intraoral Scan 
x 
 
 
x 
 
x 
Photograph 
x 
x 
x 
x 
x 
x 
Clinical 
assessment of ridge 
dimensions 
x 
x 
 
x 
 
 
Radiographic 
assessment of ridge 
dimensions 
 
x 
 
x 
 
 
Histologic 
evaluation 
 
 
 
x 
 
 
Adverse events 
recorded 
 
 
x 
 
x 
x 
Pt. satisfaction 
survey 
 
 
 
 
 
x 
Statistical 
Analysis/Final 
Report 
 
 
 
 
 
x 
 
3.2.Visual Analog Scale (VAS) survey is a questionnaire regarding post-operative pain, 
swelling, bruising effect, and daily activities tolerance. For each question, patients are 
asked to mark on a 0 to 10 scale their perception of the level of corresponding pain, 
swelling, bruising, and alterations to daily activities with 0 being no symptoms and 10 
being the worst symptom imaginable. It will be completed two weeks after every 
surgical procedure. 

 
 
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3.3.Outcomes measurements:  
 
• Clinical and Radiographic Measurements:  
Clinical measurements will be taken by one experienced blinded examiner to the randomization 
process before ridge preservation procedure and 3 months after ridge preservation, using 
intraoral scan and direct clinical assessment using a calibrated surgical caliper and UNC-15 
periodontal probe (in millimeters). Another blinded examiner will perform the radiographic 
measurements at time of ridge preservation procedure and 3 months after ridge preservation, 
using a CBCT and implant planning software (in millimeters). Horizontal and vertical ridge 
dimensions will be evaluated clinically and radiographic. Intra-examiner calibration will be 
conducted to ensure reliability of measuring method.  
• Histomorphometric Analysis: 
Each specimen will be fixated with 10% neutral buffered formalin for 48 hours prior to be 
dehydrated and embedded in methylmethacrylate. It will be ground sectioned at the center of the 
biopsy in its long axis into 50-70 micron-thick sections (Exakt Technologies, Inc., Oklahoma 
City, OK), and polished with 4000 grit sandpaper and Novus Polish to create a surface as smooth 
as possible. All sections will be stained with Sanderson’s Bone Stain and imaged for 
quantification of bone formation. Histomorphometry will be done using the Bioquant Image 
Analysis Software (R&M Biometrics, Nashville, TN) by measuring the total surface of vital 
bone, residual graft particles, organic matrix and artifact/air components. Corresponding 
percentages will be calculated for each of these tissues and compared between the three groups 
(L-PRF, L-PRF/FDBA, LPRF/FDBA layered technique). These experiments and measurements 
will be conducted by an experienced blinded lab technician.   
3.4.Statistical Analysis:  
A power analysis was conducted to determine the sufficient number of patients in each treatment 
group to obtain a power of 80% to detect a 1.0mm change in ridge dimensions using a type 1 
(alpha) error rate of 0.05. The power analysis determined that a minimum of 10 patients would 
be required for each group, and 30 patients in total. An alpha of 0.05 was assumed after a 
Bonferroni correction for five relevant 2-way comparisons between the treatment groups. 
Two-tailed t test will be used to compare clinical characteristics between patients with the L-
PRF/FDBA layered technique, L-PRF/FDBA, and those with L-PRF alone for continuous 
variables with a normal distribution. Nonparametric (Wilcoxon) test will be used otherwise.  
For categorical variables, inter-group comparisons will be conducted using chi-square test or 
Fisher’s Extract Test, as deemed necessary. To determine whether the groups experienced 
equivalent changes in bone growth from baseline to follow-up, paired t (or Wilcoxon signed-
rank) and Mc Nemar’s tests will be used for continuous and categorical variables, respectively.  
The α level will be set at 0.05. Statistical analysis will be carried out using SAS software version 
9.4 (SAS Institute, Inc, Cary, North Carolina).  

 
 
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4.0 Adverse Event Reporting 
 
Participants will be interviewed and instructed to call if they experience any adverse events after 
study procedures. Such events will be recorded. In the event of a serious adverse event, the IRB 
will be notified per posted requirements and in full compliance with federal guidelines for 
research in human subjects. 
 
5.0 Cost/Benefits to participants 
 
5.1 Cost/Benefits to participants 
There is no additional cost for study participation and study participants will pay for a reduced 
fee for the surgical procedures including extraction, grafting, and implant placement ($1200). 
This represents a fee reduction of approximately $800. Additionally, participants will receive a 
dental cleaning at no charge at the 6-month follow-up visit. 
Benefits of this project for individual participants could include enhanced bone healing, due to 
the potential benefit of L-PRF at extraction socket grafting sites. Overall benefits include the 
assessment of the practicality and utility of utilizing L-PRF either alone or in combination with a 
bone allograft in an extraction socket model to improve new bone formation and produce a more 
ideal environment for the installation of dental implant(s) at an edentulous site. 
 
 
5.2 Additional Dental Procedures 
Study visits are not designed to take the place of routine dental care. Costs for non-study related 
treatment will be charged to participant account or insurance provider. Implant restorative 
treatments including implant abutment(s), crown(s), and/or other restorative components are not 
included in the reduced surgical fee for study participants.