Clinical and Histological Outcomes of Allogenic Amnion Chorion Membrane in the Healing of Free Gingival Graft Donor Site
Phase: NA
Conditions: Palatal Wound
Sponsor: The University of Texas Health Science Center, Houston

IRB NUMBER: HSC-DB-18-0303 
IRB APPROVAL DATE: 04/03/2020 
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Protocol Number, Version No and Date 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Clinical and Histological Outcomes of Allogenic Amnion Chorion Membrane in 
the Healing of Free Gingival Graft Donor Site 
 
NCT03713073 
 
Version Date: 04/03/2020 
 
 

IRB NUMBER: HSC-DB-18-0303 
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STUDY PROTOCOL 
 
PROTOCOL NUMBER: 
HSC-DB-18-0303 
 
PROTOCOL TITLE: 
Clinical and Histological Outcomes of Allogenic Amnion Chorion Membrane in the 
Healing of Free Gingival Graft Donor Site 
VERSION: 
1 
DATE: 
02.20.2018 
CORRESPONDING PRINCIPAL INVESTIGATOR: 
Chun-Teh Lee, Assistant Professor, UTHealth School of Dentistry 
 
SITE PRINCIPAL INVESTIGATOR: 
Chun-Teh Lee, Assistant Professor, UTHealth School of Dentistry 
Seonghong Min, Assistant Professor, UTHealth School of Dentistry 
SITE: 
1. UTHealth School of Dentistry, 7500 Cambridge street, Houston, TX 77054 
 
 
COLLABORATORS: 
Nikola Angelov, Professor, UTHealth School of Dentistry 
Robin Weltman, Professor, UTHealth School of Dentistry 
Bing-Yan Wang, Professor, UTHealth School of Dentistry 

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GENERAL INFORMATION 
 
 
Name and address of the sponsor of the study: 
Not available 
 
 
 
Name and address of the person authorized to sign the protocol and 
amendments: Chun-Teh Lee, 7500 Cambridge Street, Suite 6422, Houston, Texas 
77054 Seonghong Min, 7500 Cambridge Street, Suite 6427, Houston, Texas, 77054 
Bing-Yan Wang, 7500 Cambridge Street, Suite 5316, Houston, Texas, 77054 
 
Name, title, address and telephone number(s) of the medical expert for the trial 
Chun-Teh Lee, 7500 Cambridge Street, Suite 6422, Houston, Texas 77054 Office: 
713.486.4384; Cell: 6263798070 
Seonghong Min, 7500 Cambridge Street, Suite 6427, Houston, Texas, 
77054 Office: 713.486.4391 
Office: 713.486.4384; Cell: 6263798070 Bing-Yan Wang, 7500 Cambridge 
Street, Suite 5316, Houston, Texas 77054 Office: 713.500.4385 
 
Name and title of the investigator(s) and sub-investigators responsible for the trial 
with address and phone number(s) 
Chun-Teh Lee, 7500 Cambridge Street, Suite 6422, Houston, Texas 77054 
Office: 713.486.4384; Cell: 6263798070 
Seonghong Min, 7500 Cambridge Street, Suite 6427, Houston, Texas, 
77054 Office: 713.486.4391 
Bing-Yan Wang, 7500 Cambridge Street, Suite 5316, Houston, Texas 
77054 Office: 713.500.4385 

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TABLE OF CONTENTS 
1. BACKGROUND AND RATIONALE ................................................................................................. 
5 
1.1.  GENERAL INTRODUCTION ............................................................................................................. 
5 
1.2. RATIONALE AND JUSTIFICATION FOR THE STUDY......................................................................... 
6 
A. 
RATIONALE FOR THE STUDY PURPOSE............................................................................................. 
7 
B. 
RATIONALE FOR DOSES SELECTED .................................................................................................. 
7 
C. 
RATIONALE FOR STUDY POPULATION ............................................................................................... 
7 
D. 
RATIONALE FOR STUDY DESIGN ....................................................................................................... 
7 
2. HYPOTHESIS AND OBJECTIVES .................................................................................................. 
8 
 
2.1. HYPOTHESIS.................................................................................................................................. 
8 
2.4. POTENTIAL RISKS AND BENEFITS: ................................................................................................ 
8 
A. 
END POINTS - EFFICACY.................................................................................................................... 
8 
B. 
END POINTS - SAFETY ....................................................................................................................... 
8 
3.1. LIST THE NUMBER OF SUBJECTS TO BE ENROLLED. ..................................................................... 
9 
3.2. 
CRITERIA FOR RECRUITMENT ....................................................................................................... 
9 
3.3. 
INCLUSION CRITERIA ..................................................................................................................... 
9 
3.4. EXCLUSION CRITERIA ................................................................................................................. 
10 
3.5. WITHDRAWAL CRITERIA .............................................................................................................. 
10 
3.6. SUBJECT REPLACEMENT ............................................................................................................ 
10 
4.  TRIAL  SCHEDULE............................................................................................................................ 
10 
5. 
STUDY DESIGN............................................................................................................................ 
10 
5.1.  SUMMARY OF STUDY DESIGN..................................................................................................... 
10 
6. METHODS AND ASSESSMENTS ................................................................................................. 
11 
6.1.  RANDOMIZATION AND BLINDING ................................................................................................. 
11 
6.2.  CONTRACEPTION AND PREGNANCY TESTING ............................................................................ 
11 
6.3.  STUDY VISITS AND PROCEDURES .............................................................................................. 
11 
7. TRIAL MATERIALS .......................................................................................................................... 
14 
7.1.  TRIAL PRODUCT (S) .................................................................................................................... 
14 
7.2.  STORAGE AND DRUG ACCOUNTABILITY ..................................................................................... 
15 
8. TREATMENT...................................................................................................................................... 
15 
8.1.  RATIONALE FOR SELECTION OF DOSE ....................................................................................... 
15 
8.2. 
STUDY DRUG FORMULATIONS ................................................. ERROR! BOOKMARK NOT DEFINED. 
8.3. 
STUDY DRUG ADMINISTRATION ............................................... ERROR! BOOKMARK NOT DEFINED. 
8.4. 
SPECIFIC RESTRICTIONS / REQUIREMENTS ............................................................................... 
15 
8.5.  BLINDING ..................................................................................................................................... 
15 
8.6. 
CONCOMITANT THERAPY ........................................................... ERROR! BOOKMARK NOT DEFINED. 
9. SAFETY MEASUREMENTS............................................................................................................ 
16 
9.1.  DEFINITIONS ................................................................................................................................ 
16 
9.2. 
COLLECTING, RECORDING AND REPORTING OF ADVERSE EVENTS ......................................... 
16 
9.3.  SAFETY MONITORING PLAN........................................................................................................ 
17 
10. 
DATA ANALYSIS ......................................................................................................................... 
17 
10.1. DATA QUALITY ASSURANCE ....................................................................................................... 
17 
10.2. DATA ENTRY AND STORAGE....................................................................................................... 
17 
11. 
SAMPLE SIZE AND STATISTICAL METHODS ..................................................................... 18 
11.1. DETERMINATION OF SAMPLE SIZE.............................................................................................. 
18 
11.2. STATISTICAL AND ANALYTICAL PLANS........................................................................................ 
18 
12. 
ETHICAL CONSIDERATIONS ................................................................................................... 
19 

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12.1. 
INFORMED CONSENT............................................................................................................................. 19 
12.2. 
IRB REVIEW ............................................................................................................................................. 20 
12.3. 
CONFIDENTIALITY OF DATA AND PATIENT RECORDS ...................................................................... 20 
13. 
PUBLICATIONS ................................................................................................................ 20 
14. 
RETENTION OF TRIAL DOCUMENTS ............................................................................ 20 

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STUDY PROTOCOL 
 
1. 
BACKGROUND AND RATIONALE 
 
 
1.1. 
General Introduction 
 
Free gingival graft (FGG) procedure is commonly conducted to increase the width of 
keratinized gingiva around teeth and implants. Although FGG procedure has 
predictable clinical outcomes, morbidities and complications on the donor sites 
(palate) usually affect patients’ willingness to receive the procedure (Griffin et al. 
2006). Several alternative materials, such as allogenic dermal matrix or xenogeneic 
collagen matrix, have been used to prevent these complications, but these 
substitutes have inferior clinical results to FGG (McGuire and Scheyer 2014; Wei et 
al. 2000). Since FGG has an irreplaceable position in augmenting keratinized gingiva, 
clinicians utilize dressings to improve palatal wound healing (Shanmugam et al. 
2010; Thoma et al. 2016). Allogenic amnion chorion membrane (ACM) has been 
used to facilitate wound healing, such as diabetic ulcer (Zelen et al. 2013), chronic 
wounds(Forbes and Fetterolf 2012) and perforated sinus membrane (Holtzclaw 2015) 
because this placental membrane retains extracelluar matrix proteins, growth factors 
and cytokines to promote cell proliferation and control inflammation at the wound 
(Koob et al. 2013). Additionally, allogenic amnion chorion membrane can be well 
attached to the wound because of its thin thickness and flexibility. Due to these 
characteristics, allogenic amnion chorion membrane might be an excellent dressing 
for palatal wound after harvesting a FGG. 
1.2. 
Rationale and justification for the Study 
 
 
a. 
Rationale for the Study Purpose 
So far, there is no strong evidence supporting the clinical impact of any dressings 
or membranes on the healing of the FGG donor site. It is very important to assess the 
effects of the allogenic amnion chorion membrane on the wound of the FGG donor 
site given this membrane has the potential to improve wound healing and reduce 
complications. 
b. 
Rationale for Materials Selected 
 
The ACM (allogenic amnion chorion membrane, BioXclude®, SNOASIS MEDICAL) 
is a commercial product and has multiple sizes available. It has been used for tissue 
healing and tissue regeneration in dentistry. Based on the available evidence, it has 
the potential to improve wound healing of the FGG donor site. The collagen dressing, 
CLD (Collagen Patch®, Zimmer), is a common product used for tissue healing in 
dentistry(Shanmugam et al. 2010). It is selected as a dressing material to cover the 
wound in the control group. 
 
c. 
Rationale for Study Population 
 
The study will recruit subjects who need free gingival graft surgery. The dressings, 
ACM or CLD, will be placed in the wound of the FGG donor site. These subjects 

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should be systemically healthy and are able to receive FGG surgery. . 
d. 
Rationale for Study Design 
 
It is a randomized controlled and split-mouth design study. The randomized 
controlled trail (RCT) is usually considered the gold standard for a clinical trial and it 
is able to provide the highest level of evidence within all the study designs (Howick et 
al. 2011). The split-mouth design is a popular design in oral health research. Two 
treatments (test and control) are randomly assigned to either the right or left halves 
of the dentition/palate. This design removes a lot of inter-individual variability from 
the estimates of the treatment effect given two treatments are performed in the same 
subject (Lesaffre et al. 2009). 
 
 
2. 
HYPOTHESIS AND OBJECTIVES 
 
 
2.1. 
Hypothesis 
 
Null hypothesis: 
Healing at the palatal wound covered with an allogenic amnion chorion 
membrane and the collagen dressing has no significant difference. Alternative 
hypothesis: 
The palatal wound covered with an allogenic amnion chorion membrane heals 
faster than the palatal wound covered with a collagen dressing. The patients with an 
allogenic amnion chorion membrane feel less pain and have fewer complications 
than the patients with a collagen dressing. 
2.2. 
Primary Objectives 
 
This study aims to compare an allogenic amnion chorion membrane (BioXclude®, 
SNOASIS MEDICAL) to a collagen dressing (Collagen Patch®, Zimmer) in the 
palatal wound healing from the clinical and histological perspectives. 
2.3. 
Secondary Objectives 
 
In addition to the outcomes of wound healing, the clinician’s feedback with 
these two materials (ACM and CLD), such as handling and operation time, will also 
be evaluated. 
2.4. 
Potential Risks and Benefits: 
 
a. 
End Points - Efficacy 
All eligible patients need FGG procedure and will receive the appropriate 
treatments. The patients may have improved healing or may not have 
additional clinical benefits by participating in this clinical study. 
b. 
End Points - Safety 
1) Study related risks: 
Initial infection, pain, swelling, and bleeding related to FGG procedure and gingival 

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biopsy; adverse outcomes of the receipt sites; The patients will have the same risks if 
they receive the same treatment without participating in the study. 
2) Protection against risks: 
All efforts will be made to minimize risks to all and every participant: only patients 
with the need for FGG procedure will qualify for the study. 
 
3. 
STUDY POPULATION 
 
3.1. 
List the number of subjects to be enrolled. 
The proposed project is a pilot study. We use a convenient sample size based on 
the budget. We will recruit 19 patients in total, and with a dropout rate of 15%, we will 
have a sample size of 15. 
With 15 patients, we can detect an effect size as small as 0.7 between two 
treatments for the two main outcomes (the area of healing and VAS pain score 
at week 2), by a paired t test assuming a within-subject correlation of 0.6 at the 
significance level of 0.05 with 80% power. 
3.2. 
Criteria for Recruitment 
 
The patients who need FGG procedure will be recruited in the study. These 
patients should be systemically healthy to receive this procedure. 
3.3. 
Inclusion Criteria 
 
Nineteen subjects will be recruited among the patients attending the Clinic for 
Graduate Periodontics, UTSD, who are in need of a FGG procedure to augment 
keratinized gingiva, augment ridge or cover recession defects. The size of the FGG 
that the patient needs will not be bigger the size of FGGs that have to be harvested in 
this study (two 8mm (width) x 10mm (length) x ≈1.5mm (thickness) FGG). All 
subjects are ≥18 year-old and systemically healthy or with controlled common 
systemic conditions, such as hypertension, that will not affect wound healing. 
3.4. 
Exclusion Criteria 
 
Patients will be excluded if they are current heavy smokers(>10 cigarettes/day), 
have diabetes or other systemic diseases that may comprise healing, take 
antibiotics and/or analgesics within one week before the procedure and have loss of 
sensation on the palate. Patients who stop smoking more than one year are eligible. 
3.5. 
Withdrawal Criteria 
 
A subject may be discontinued from participation in the study for any of 
the following reasons: 
1. Withdrawal of consent 
2. Subject noncompliance with the protocol, as determined by the investigator 
3. Any event or condition that would make continued participation in the study not in 
the best interest of the subject, as determined by the investigator 
4. Pregnancy 
5. Development of any medical condition that might affect the treatment and clinical 
outcomes, as determined by the investigator. 
6. Initiation of any treatment or exposure that might affect the healing outcomes of 
the FGG donor site, as determined by the investigator. 

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7. Investigator discretion 
 
3.6. 
Subject Replacement 
 
Subjects who withdraw from the study can be replaced. However, to complete the 
study within the time allocated, the center will not enroll subjects after 24 months from 
enrollment initiation. 
 
4. 
TRIAL SCHEDULE 
 
There will be eight appointments including the baseline appointment and seven 
follow-up appointments (4 days, 10 days, 2, 3, 4, 6, 8 weeks) after the surgery. The 
details of each visit will be mentioned in 6.3. Study Visits and Procedures. 
 
5. 
STUDY DESIGN 
 
 
5.1. 
Summary of Study Design 
 
This randomized controlled and split-mouth design study aims to compare an 
allogenic amnion chorion membrane (BioXclude®, SNOASIS MEDICAL) to a 
collagen dressing (Collagen Patch®, Zimmer) in the palatal wound healing from the 
clinical and histological perspectives. Nineteen patients will be recruited. The wound 
healing would be assessed using pictures and questionnaires at multiple time points. 
A biopsy will be done to evaluate histological healing. Patients will not have 
additional complications other than the complications from FGG procedure. However, 
the size of wounds will be larger than the size required to treat defects. The 
hypothesis is that the patients with an allogenic amnion chorion membrane feel less 
pain and have fewer complications than the patients with a collagen matrix. 
The potential application of the allogenic amnion chorion membrane in palatal wound 
will benefit patients who receive FGG procedure. The results can be applied in 
managing various wounds in the oral cavity in the future. 
 
 
6. 
METHODS AND ASSESSMENTS 
 
 
6.1. 
Randomization and Blinding 
 
Site allocation (right or left side of the palate) to the two groups (ACM or CLD) 
will be performed by the investigators before the surgeon harvests FGGs, based on 
computer-generated randomization (R Statistical Software). Patients will be blinded 
because they will not know which dressing is in the test group or the control group. 
6.2. 
Contraception and Pregnancy Testing 
 
Pregnancy status of subjects who are women with childbearing potential will be 
orally confirmed at the screening. The pregnant subjects will be excluded from the 
study. 

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6.3. 
Study Visits and Procedures 
 
. 
 
a. Screening Visits and Procedures 
 
Study protocol and consent forms will be approved by the Institutional Review 
Board at the University of Texas Health Science Center at Houston. The trial will be 
registered with ClinicalTrials.gov. The clinicians in UTSD will be told the information 
of this clinical trial. The potential subjects will be identified in the clinic of 
Department of Periodontics, for initial screening. The principal investigator will 
confirm the eligibility of these patients. 
All patients will sign the consent forms and are informed of the details of study 
procedures as well as potential complications. After informed consent is obtained, 
the surgery will be scheduled as the first visit. An impression will be taken to make 
the stent before the surgery. 
 
b. Study Visits and Procedures 
 
Patients will have FGGs harvested from two sites on the palate (right and left; the 
area between canine to first molar). In addition to the FGG wounds, two palatal 
gingiva biopsies will be harvested. The size of the FGG will be standardized (8mm 
(width) x 10mm (length) x ≈1.5mm thickness). FGGs will be harvested by the #15 
blade with the assistance of a template stent. Two FGGs will be used based on the 
patient’s need. If the patients need grafts larger than two standardized FGGs in one 
procedure, the patients will be excluded from the study. The biopsies of palatal 
gingiva will be harvested by a tissue punch and the size is standardized (4mm 
(diameter) x ≈1.5mm thickness). The area of biopsy will be at least 3mm away from 
the FGG wound. Biopsies and FGGs will be harvested in the area between maxillary 
canine and first molar where tissue grafts are commonly harvested from (Reiser et 
al. 1996). These wounds can be completely healed even without placing any 
dressing materials (Harris et al. 2007). Dressing materials are used to improve 
healing and reduce complications. The wounds of FGG and biopsy will be covered 
with an allogenic amnion chorion membrane (ACM, BioXclude®) or a collagen 
dressing (CLD, Collagen Patch®) based on the randomization table. 
Clinical Measurements 
 
The donor sites of FGG will be evaluated on 4 days, 10 days, 2 weeks, 3 weeks, 4 
weeks, 6 weeks, 8 weeks after the surgery (7 follow-up visits in total). Healing of the 
donor site will be analyzed based on (1) the degree of epithelialization, (2) the size of 
wound area, (3) color match and (4) bleeding condition. 
(1) Epithelization will be evaluated at each follow-up visit until the wound is 
completely epithelized. It is measured by means of bubble formation after dripping 
hydrogen peroxide (3%) to the wound surface and epithelization will be ranked as 
total, partial or none (Keceli et al. 2015; Marucha et al. 1998); 
(2) The wound area will be assessed by means of a clinical picture taken at each 
follow-up visit (Del Pizzo et al. 2002; Thoma et al. 2012). The picture will be taken 
with an angulation perpendicular to the wound. A periodontal probe will be placed 
by the wound as a reference scale. Wound area is defined as the area without 

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epithelization. The wound area on the picture will be measured by ImageJ; 
(3) Color match of the donor site will be identified at every visit by using a 4-point scale: 
1. Obvious difference; 2. Noticeable difference; 3. Disguisable difference; 4 No 
noticeable difference in comparison with adjacent gingiva. Three measurements, (1), 
(2), and (3), will be performed by one independent examiner who does not know the 
dressing (ACM or CLD) for each wound; 
(4) Haemostasis has to be achieved when no bleeding is actively seen. Delayed 
bleeding, prolonged haemorrhaging from the palate during the postsurgical period, 
reported by the patient, will be documented (Del Pizzo et al. 2002). 
The wound healing will also be evaluated from the patient’s perspective by means of 
(5) pain and (6) sensibility. Patients will be given a questionnaire to answer these 
questions every day or at the scheduled appointments (4 days, 10 days, 2 weeks, 
3 weeks, 4 weeks, 6 weeks, 8 weeks) until the patient completes the study. 
(5) Pain will be assessed by using VAS pain scores (0 to 10. 0: no pain, 1: 
minimal pain, 10: severe pain) (Keceli et al. 2015); 
(6) Sensitivity will be assessed by means of a periodontal probe (XP 23/UNC 15, Hu- 
Friedy) using a 4-point discrimination scale (coronal, apical, mesial, distal) around the 
donor area, before the surgery and at the follow-up visits. Sensibility will be recorded 
using a rubbing movement and a pin-pressure nociception. Patients will be asked to 
give a rating of their loss of sensibility based on a three-point verbal descriptor scale 
(VDS): none, mild or moderate, severe) (Del Pizzo et al. 2002); 
(7) The surgeons will be given a questionnaire to answer questions 
regarding handling of the two different materials after performing the surgery. 
Volumetric Analysis 
The impression of FGG donor site will be obtained using intraoral scanner at pre and 
post-operatively 4 days, 10 days, 2 weeks, 3 weeks, 4 weeks, 6 weeks, 8 weeks. 
The STL files of pre-op and post-op will be imported into the reverse engineering 
software. Then, three-dimensional images will be reconstructed and superimposed to 
allow for the measurement of volumetric change within the initial harvested area 
during wound healing phase. 10% of the sites will be randomly selected for repeated 
measurements. 
Histological and Histomorphometric Analyses 
Gingival specimens (4mm in diameter, 1.5mm in thickness) will be harvested at 
baseline visit and the 10-day follow-up visit for histological and histomorphometric 
analyses. The second biopsy will be performed at the 10-day follow-up visit because 
epithelization will not be completed before two weeks. The tenth day is a proper 
timing to evaluate early healing. Specimens will be cut in sections (5µm in thickness) 
and stained with either H&E to identify the cellular composition of inflammatory 
infiltrates or Masson’s trichrome to detect new collagen depositions. For qualitative 
histological analysis, the histological sections will be evaluated using a Nikon light 
microscope, using Nikon NIS Element AR Imagine Software (Nikon Metrology, Inc. 
Brighton, MI) to calculate the percentages of the new collagen and matured collagen, 
as well as the thickness of the epithelium automatically. Three regions of interest will 
be analyzed on each specimen corresponding to the superficial, central, and apical 
third. Immunohistochemical staining will be performed to detect factor VIII (von 
Willebrandt factor) to detect endothelial cells and therefore angiogenesis. The area of 
angiogenesis will be quantified using the same Nikon NIS Element AR Imagine 
Software. 

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c. Final Study Visit: 
 
The 8-week follow-up will be the subject’s last visit for this clinical trial. 
The patients will have clinical measurements as previously mentioned and 
received the gift card. 
d. Post Study Follow up and Procedures 
 
The subjects will continue having routine maintenance appointments to follow up 
the outcomes related to the surgery and periodontal health in the periodontics clinic 
of UTSD. If the patients have symptoms or complications, the necessary treatments, 
will be performed. 
e. Discontinuation Visit and Procedures 
 
Subjects are free to withdraw from participation in the study at any time upon 
request. A subject may be discontinued from participation in the study for any of 
the following reasons: 
1. Withdrawal of consent 
2. Subject noncompliance with the protocol, as determined by the investigator 
3. Any event or condition that would make continued participation in the study not 
in the best interest of the subject, as determined by the investigator 
4. Pregnancy 
5. Development of any medical condition that might affect the treatment and clinical 
outcomes, as determined by the investigator. 
6. Initiation of any treatment or exposure that might affect the outcomes of 
implant therapy, as determined by the investigator. 
7. Investigator discretion 
Any subject with a serious adverse event, such as life-threating diseases, 
hospitalization, that is ongoing at the time of discontinuation will be followed until 
the event returns to baseline, resolves, or stabilizes. If the serious adverse event 
does not meet these outcomes within 30 days after discontinuation or after study 
completion, the subject will be referred to an appropriate practitioner for continued 
care. If the study is discontinued, subjects will be referred back to the qualified 
clinicians for necessary dental care. 
 
 
7. 
TRIAL MATERIALS 
 
. 
 
7.1. 
Trial Product (s) 
 
Allogenic amnion chorion membrane (ACM), BioXclude, is a minimally manipulated 
allograft amnion chorion tissue for use as a wound covering in dental surgery. It has 
been widely used in dentistry. Its unique physical and biological properties provide the 
benefits of a growth factor and an occlusive barrier. The tissue used in BioXclude is 
obtained from consenting mothers who donate their placentas after elective caesarian 
section delivery. The amnion chorion tissue is processed using a patented tissue 
processing technology (Purion®,) designed to cleanse and maintain the delicate 
structures of the tissue. Following processing and dehydration, the allografts are 
packaged and terminally sterilized. This processing methodology allows for 

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retention of the biological factors found in native amnion chorion tissue. 
 
Collagen dressing (Collagen Patch®, Zimmer) is used to cover wound in dental 
surgery. It can adhere to the wound and retain its structural integrity even when 
wet. It is known to control bleeding and stabilize blood clots as well as protect the 
wound bed while accelerating the healing process. 
7.2. 
Storage and Drug Accountability 
 
BioXclude membrane and Collagen dressing will be stored at ambient 
temperature in its original packaging following the manual instruction. The expiration 
date for the product is recorded on the product container labeling as year (4 digits) 
and month (2 digits) and the product expires on the last day of the month indicated. 
Expiration date printed on the labeling is valid as long as product is stored at 
ambient temperature and in an unopened foil pouch/packaging. Once the product is 
expired, the material will be discarded. 
 
8. 
TREATMENT 
8.1. 
Rationale for Selection of Treatments 
 
All subjects need FGG procedure and FGGs are harvested for clinical and research 
purposes. The treatments are conducted following general clinical principles. The 
details of procedure are mentioned in section 6.3. 
8.2. 
Specific Restrictions / Requirements 
 
 
Patients will be prescribed an analgesic (Ibuprofen, 600 mg, q.i.d; if the patient 
cannot take ibuprofen, 500mg acetaminophen, q.i.d will be prescribed). A disinfectant 
solution (Chlorhexidine digluconate, 0.2% solution) will be prescribed but the patient 
will only apply the solution to the recipient site using a monojet syringe (q 8 hours, 10 
days). An individualized protective stent will be given to the patient. The patient will 
be instructed to wear the stent overnight after the surgery and continue wearing it for 
10 days. The patient will be instructed to take liquid diet for the first 2 days then soft 
diet till the 10th day. Sutures at the FGG site will be removed 4 days after surgery. 
 
8.3. 
Blinding 
 
The surgeons cannot be blinded because the surgeons will be told which dressing 
should be placed on the specific site. Patients will be blinded given they will not know 
how the surgical group will be assigned. The patients will be informed of benefits of all 
the procedures and realize all procedures are supported by scientific evidence. 
 
 
9. 
SAFETY MEASUREMENTS 
9.1. 
Definitions 
All unanticipated problems will be reported in this study. The Committee For the 
Protection of Human Subjects (CPHS) considers unanticipated problems to be any 

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incident, experience, or outcome that meets all of the following criteria: 
Is unexpected in terms of nature, severity, or frequency given a) the research 
procedures that are described in the IRB-approved research protocol and informed 
consent, and b) the characteristics of the subject population being studied; 
Is related or possibly related to participation in the research (possibly related 
means there is a reasonable possibility that the incident, experience, or outcome 
may have been caused by the procedures involved in the research); and 
Places subjects or others at a greater risk for physical, psychological, economic, 
or social harm than was previously known or recognized. 
An incident, experience, or outcome that meets the 3 criteria above will generally 
warrant consideration of substantive changes in order to protect the safety, welfare, or 
rights of subjects or others. Examples of corrective actions or substantive changes that 
might need to be considered in response to an unanticipated problem include the 
following: 
Changes to the research protocol initiated by the investigator prior to obtaining 
IRB approval to eliminate apparent immediate hazards to subjects 
Modification of inclusion or exclusion criteria to mitigate newly identified 
risks Implementation of additional procedures for monitoring subjects 
Suspension of enrollment of new subjects 
Suspension of research procedures in currently enrolled subjects 
 
For this study, an severe adverse event (SAE) is defined as an unanticipated 
problem occurring during the study that fulfils 1 or more of the following criteria: 
1. Results in death 
2. Is immediately life-threatening† 
3. Requires inpatient hospitalization or prolongation of existing hospitalization 
4. Results in persistent or significant disability or incapacity 
5. Is a congenital abnormality or birth defect 
6. Is an important medical event that may jeopardize the subject or may 
require medical intervention to prevent one of the outcomes listed above 
Hospitalization for elective procedures or surgeries will not be considered SAEs, 
nor will inpatient hospitalizations for convenience. 
Pregnancy in women with childbearing potential should not be reported as an 
SAE, but if pregnancy occurs, it must be reported in accordance with the 
procedures described in Section 6.2. Pregnancy will not be regarded as an SAE 
unless there is suspicion that a study intervention may have interfered with the 
effectiveness of a contraceptive medication and the event meets the criteria for an 
unanticipated problem. If the pregnancy results in an outcome other than a normal 
birth or elective abortion of a healthy fetus, it will be reported as an SAE. 
9.2. 
Collecting, Recording and Reporting of Adverse Events 
 
Examination and close follow-up of parameters capturing subjects’ oral health will 
be collected on case report forms (CRFs). These will be completed at every study 
visit, and data will be compiled into a pre-specified format and reviewed monthly by 
the PI for safety oversight. 
Serious adverse events (as defined in Section 9.1) will be collected from the time 
of enrollment until the last clinic visit and will be recorded in the electronic health 
records (EHR) system. At each study visit, the clinician or investigator will inquire 
about the occurrence of SAEs since the last assessment. The investigator will review 
all source documentation related to study procedures for evidence of SAEs. Events 
will be followed for outcome information until they return to baseline or stabilize, or 

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until 30 days after study completion or subject discontinuation. Subjects who have an 
SAE that is ongoing 30 days after study completion or discontinuation will be referred 
to an appropriate practitioner for continued care. 
Upon learning that a subject has experienced an SAE, the investigator must report 
the event to CPHS within 24 hours after becoming aware of the event. 
On a monthly basis, the following events will be reported to every PI: 
Number of subjects experience severe complications and number of subjects 
enrolled. Severe complications include severe pain (VAS>6), continuous bleeding 
and severe swelling that needs prescription to control. 
Duration of observation of subjects experiencing severe complications and 
duration of observation of subjects enrolled. 
Any tooth loss, abscess, or other adverse oral health development requiring 
therapy or other intervention and the etiology (as captured in the dental history) 
Every PI will review the monthly reports for any safety signals. 
9.3. 
Safety Monitoring Plan 
 
The purposes of the clinical monitoring activities are to ensure that the rights of 
human subjects are protected, the study is implemented in accordance with the 
protocol, and the integrity of study data is maintained. 
All subjects will be monitored for postoperative healing and tissue response at a 
regular interval while the entire oral health will be maintained throughout the 
study period. 
 
 
10. 
DATA ANALYSIS 
10.1. Data Quality Assurance 
 
Data and measurements will be checked by two separate investigators (Chun- 
Teh Lee, Seonghong Min) as well as analyzed statistically to ensure that the data 
obtained is accurate, complete and reliable. 
10.2. Data Entry and Storage 
 
Case report forms (CRFs) will be completed and stored in a locked file cabinet in the 
PI’s office located at UTSD. Data will be entered electronically in excel spreadsheets, 
and images will be stored electronically; both will be stored on the PIs work computer 
in a locked office and password protected. 
 
 
11. 
SAMPLE SIZE AND STATISTICAL METHODS 
 
11.1. Determination of Sample Size 
 
The pilot project will use a convenient sample size based on the budget. We will 
recruit 19 patients in total, and with a dropout rate of 15%, we will have a sample size 
of 15. 
With 15 patients, we can detect an effect size as small as 0.7 between two 
treatments for the two main outcomes (the area of healing and VAS pain score at 

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week 2), by a paired t test assuming a within-subject correlation of 0.6 at the 
significance level of 0.05 with 80% power. Considering this is a pilot study, 
Bonferroni correction is not necessary. 
 
11.2. Statistical and Analytical Plans 
a. General Considerations 
 
We will present means and standard deviations for completeness of the report. The 
statistical significance level to test the primary endpoint was set at p<0.05, a priori. 
b. Safety Analyses 
 
Safety will be evaluated by tabulations of adverse events and will be presented 
with descriptive statistics at baseline and follow-up visits each month. 
Adverse events will be classified as severe complications and summarized 
for baseline and follow-up visits. 
All information pertaining to adverse events noted during the study will be listed by 
subject, detailing verbatim given by the investigator, preferred term, date of onset, 
date of resolution, severity, and relationship to procedure. The onset of adverse 
events will also be shown relative (in number of days) to the day of performing the 
surgery. 
A tabulation of Serious Adverse Events (SAEs) will be provided by subject within 
treatment groups. The proportion of subjects in each treatment group reporting 
adverse events that occur in ~ 3% in either treatment group will be compared using 
Bayesian methods. The specific preferred terms analysed will be those that are 
reported by at least five percent of the subjects in either treatment group. 
 
c. Statistical Analysis Plan: 
We will explore the difference between the two treatments for outcomes including 
the degree of epithelialization, the size of wound area, color match, bleeding 
condition, pain, sensibility, and outcomes from Gingival specimens by paired t test or 
McNemar's test depending on the variables. 
 
12. 
ETHICAL CONSIDERATIONS 
12.1. Informed Consent 
 
Informed consent is a process that is initiated prior to the individual’s agreeing to 
participate in the study and continues throughout the individual’s study participation. 
Extensive discussion of risks and possible benefits of study participation will be 
provided to the subjects and their families. A consent form describing in detail the 
study interventions, procedures, and risks will be given to the subject. Consent forms 
will be IRB-approved, and the subject will be asked to read and review the document. 
The investigator or designee will explain the research study to the subject and 
answer any questions that may arise. The subject will sign the informed consent 
document prior to any study-related assessments or procedures. Subjects will be 
given the opportunity to discuss the study with their surrogates or think about it prior 

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to agreeing to participate. They may withdraw consent at any time throughout the 
course of the trial. A copy of the signed informed consent document will be given to 
subjects for their records. The rights and welfare of the subjects will be protected by 
emphasizing to them that the quality of their clinical care will not be adversely 
affected if they decline to participate in this study. 
The consent process will be documented in the clinical or research record. 
This documentation will include the following: 
A notation of the date that the consent was obtained 
A statement that the consent was obtained prior to the initiation of study 
procedures 
A statement that the subject had adequate time to review the consent and that 
all questions were answered prior to initiation of study procedures 
A notation confirming that a copy of the signed consent was given to the subject 
 
12.2. IRB review 
 
The protocol, informed consent form(s), and all advertising and subject materials will 
be submitted to the IRB for review and approval. Approval of both the protocol and 
informed consent form must be obtained before the enrollment of any subject. Any 
amendment to the protocol will require review and approval by the IRB before the 
changes are implemented in the clinic. 
12.3. Confidentiality of Data and Patient Records 
 
The subject’s name will appear only on the consent form and clinical record, both of 
which will be kept separate from collected study data. All subject files will be kept 
confidential and placed in a double-locked office. A unique coded study number will 
be assigned to each subject for data collection. The number will not contain any 
personal information (e.g., dates, age) to further ensure protection. 
The study protocol, documentation, data, and all other information generated will be 
held in strict confidence. No information concerning the study or the data will be 
released to any unauthorized third party without prior written approval of the PI. No 
subject names will be used in publications or presentations. 
 
13. 
PUBLICATIONS 
 
Following completion of the study, the investigator is expected to publish the results 
of this research in a scientific journal. The International Committee of Medical Journal 
Editors (ICMJE) member journals have adopted a trials-registration policy as a 
condition for publication. This policy requires that all clinical trials be registered in a 
public trials registry such as ClinicalTrials.gov (De Angelis et al. 2004), which is 
sponsored by the National Library of Medicine. Other biomedical journals are 
considering adopting similar policies. For grants and cooperative agreements, it is 
the institution’s responsibility to register the trial in an acceptable registry. 
The ICMJE defines a clinical trial as any research project that prospectively assigns 
human subjects to intervention or comparison groups to study the cause-and-effect 
relationship between a medical intervention and a health outcome. Studies designed 
for other purposes, such as to study pharmacokinetics or major toxicity (e.g., phase I 
trials), would be exempt from registering trials in a public registry such as 
ClinicalTrials.gov. 

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14. 
RETENTION OF TRIAL DOCUMENTS 
 
Patients will be assigned identifying codes that will be linked to all collected study data, 
stored in secured database by PI. All the electronic files will be encrypted and are stored 
in primary investigator's external drive, that will be locked in the PI's office cabinet. Stents 
will be stored in a locked cabinet in the PI's office. The following individuals/ institutions 
will have access to the records: the Principal Investigator and coinvestigators, and the 
University of Texas Health Science Center at Houston, including the Institutional Review 
Board. Absolute confidentiality cannot be guaranteed because of potential need to share 
this information with the above parties. The aggregate results of this study, with 
preservation of patient confidentiality, may be used for teaching, meeting presentation or 
publishing purpose. Records will be maintained for at least 6 years from the starting date 
of each subject. 
 
 
 
 
 
 
 
List of Possible Attachments 
Appendix 1 
Case report form 
Appendix 2 
Patient report form 
Appendix 3 
Flyer 
Appendix 4 
Informed Consent Form 
Appendix 5 
Schematic of Study Design 
Appendix 6 
Study Schedule 
 
 
 
References: 
Benavides E, Rios HF, Ganz SD, An CH, Resnik R, Reardon GT, Feldman SJ, Mah 
JK, Hatcher D, Kim MJ et al. 2012. Use of cone beam computed tomography in 
implant dentistry: The international congress of oral implantologists consensus 
report. Implant dentistry. 21(2):78-86. 
De Angelis C, Drazen JM, Frizelle FA, Haug C, Hoey J, Horton R, Kotzin S, Laine C, 
Marusic A, Overbeke AJ et al. 2004. Clinical trial registration: A statement from 
the international committee of medical journal editors. The New England journal 
of medicine. 351(12):1250-1251. 
Del Pizzo M, Modica F, Bethaz N, Priotto P, Romagnoli R. 2002. The connective tissue 
graft: A comparative clinical evaluation of wound healing at the palatal donor site. 
A preliminary study. Journal of clinical periodontology. 29(9):848-854. 

IRB NUMBER: HSC-DB-18-0303 
IRB APPROVAL DATE: 04/03/2020 
Page 18 of 19 
Protocol Number, Version No and Date 
 
Forbes J, Fetterolf DE. 2012. Dehydrated amniotic membrane allografts for the 
treatment of chronic wounds: A case series. Journal of wound care. 21(6):290, 
292, 294-296. 
Griffin TJ, Cheung WS, Zavras AI, Damoulis PD. 2006. Postoperative 
complications following gingival augmentation procedures. Journal of 
periodontology. 77(12):2070-2079. 
Harris RJ, Harris LE, Harris CR, Harris AJ. 2007. Evaluation of root coverage with two 
connective tissue grafts obtained from the same location. The International 
journal of periodontics & restorative dentistry. 27(4):333-339. 
Holtzclaw D. 2015. Maxillary sinus membrane repair with amnion-chorion barriers: 
A retrospective case series. Journal of periodontology. 86(8):936-940. 
Howick J, Chalmers I, Glasziou P, Greenhalgh T, Heneghan C, Liberati A, Moschetti I, 
Phillips B, Thornto H. 2011. Explanation of the 2011 oxford centre for evidence- 
based medicine (ocebm) levels of evidence (background document). 
Keceli HG, Aylikci BU, Koseoglu S, Dolgun A. 2015. Evaluation of palatal donor 
site haemostasis and wound healing after free gingival graft surgery. Journal 
of clinical periodontology. 42(6):582-589. 
Koob TJ, Rennert R, Zabek N, Massee M, Lim JJ, Temenoff JS, Li WW, Gurtner G. 
2013. Biological properties of dehydrated human amnion/chorion composite 
graft: Implications for chronic wound healing. International wound journal. 
10(5):493-500. 
Lesaffre E, Philstrom B, Needleman I, Worthington H. 2009. The design and analysis of 
split-mouth studies: What statisticians and clinicians should know. Statistics in 
medicine. 28(28):3470-3482. 
Marucha PT, Kiecolt-Glaser JK, Favagehi M. 1998. Mucosal wound healing is 
impaired by examination stress. Psychosomatic medicine. 60(3):362-365. 
McGuire MK, Scheyer ET. 2014. Randomized, controlled clinical trial to evaluate a 
xenogeneic collagen matrix as an alternative to free gingival grafting for oral 
soft tissue augmentation. Journal of periodontology. 85(10):1333-1341. 
Reiser GM, Bruno JF, Mahan PE, Larkin LH. 1996. The subepithelial connective tissue 
graft palatal donor site: Anatomic considerations for surgeons. The International 
journal of periodontics & restorative dentistry. 16(2):130-137. 
Shanmugam M, Kumar TS, Arun KV, Arun R, Karthik SJ. 2010. Clinical and 
histological evaluation of two dressing materials in the healing of palatal 
wounds. Journal of Indian Society of Periodontology. 14(4):241-244. 
Thoma DS, Hilbe M, Bienz SP, Sancho-Puchades M, Hammerle CH, Jung RE. 2016. 
Palatal wound healing using a xenogeneic collagen matrix - histological outcomes 
of a randomized controlled clinical trial. Journal of clinical periodontology. 
43(12):1124-1131. 
Thoma DS, Sancho-Puchades M, Ettlin DA, Hammerle CH, Jung RE. 2012. Impact of a 
collagen matrix on early healing, aesthetics and patient morbidity in oral mucosal 
wounds - a randomized study in humans. Journal of clinical periodontology. 
39(2):157-165. 
Wei PC, Laurell L, Geivelis M, Lingen MW, Maddalozzo D. 2000. Acellular dermal 
matrix allografts to achieve increased attached gingiva. Part 1. A clinical 
study. Journal of periodontology. 71(8):1297-1305. 

IRB NUMBER: HSC-DB-18-0303 
IRB APPROVAL DATE: 04/03/2020 
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Protocol Number, Version No and Date 
 
Zelen CM, Serena TE, Denoziere G, Fetterolf DE. 2013. A prospective randomised 
comparative parallel study of amniotic membrane wound graft in the 
management of diabetic foot ulcers. International wound journal. 10(5):502-507.