RAJIVGANDHIUNIVERSITY OF HEALTH SCIENCES
BANGALORE, KARNATAKA
ANNEXURE – II
Proforma for Registration of Subjects for Dissertation
1 / Name and Address of the Candidate / Dr. A Jagadeesh KumarPostgraduate Student
Dept. of Oral and Maxillofacial Pathology
AECSMaarutiCollege of Dental Sciences and Research Centre,
Bangalore – 560 076
2 / Name of the Institution / AECSMaarutiCollege of Dental Sciences and Research Centre
3 / Course of Study and Subject / Master of Dental Surgery in
Oral and Maxillofacial Pathology
4 / Date of Admission to the Course / June 2013
5 / Title of the Topic / Reliability of sex determination from pulp tissue using fluorescent body test.
6. Brief resume of intended work
Introduction:
Human identification is the mainstay of civilization and identification of unknown individuals has always been of paramount importance to society 1.It may require simple procedures such as logging into a computer network or more complex situations like post-mortem identification and criminal analysis. The most reliable means of identification include fingerprints, photographs, iris patterns, dental comparisons, and biological methods such as DNA profiling. 2
Forensic identification over a period of time has evolved into an art of science and involves various specialities. A doctor’s role as forensic pathologist and/or a forensic odontologist goes hand in hand with the police officer in establishing the “IDENTITY” of an individual in mass disasters. Teeth are the most durable organs in the body and can be heated to temperatures of 1,6000C without appreciable loss of microstructure. Teeth can survive even long after the soft and skeletal tissues of the body get destroyed. Biological evidence generally means the comparison of genetic material like DNA among different individuals, but DNA analysis can be expensive and time consuming.3
Dental identificationhas long been considered a reliable method when other methods fail because of critical body conditions or unavailability of body parts. Tooth pulp can be embedded in hard tissue casting that protects it from the detrimental effects of trauma and heat. So pulp can be used as a source for identification of gender of the person as sex determination is the first step in forensic identification.
When traffic accidents, acts of terrorism or mass disasters like fire accidents, tsunami, earthquake occur in which it is difficult to identify a person according to fingerprints or dental records, dental pulp may become a helpful source for personal identification. 2
6.1. Need for the study
The prime objective of the present study is to ascertain whether it is reliable to determine sex of an individual from pulp tissue using fluorescent body test.
6.2. Review of literature
1.Yamada Y et al(1989)conducted a study on extracted DNA using Southern blot hybridization for "Myo" and a Y chromosome-specific DNA probe and suggested that DNA extracted from dental pulp is useful for identification of individuals, paternity testing, and sex determination.10
2.Adachi H. (1989)conducted a study to calculate the appearance rate of Y chromatin by using Quinacrine mustard stain on the dental pulp of extracted teeth left at the room temperature. The morphological findings of the tissue were observed with the H&E stained section and the change of the appearance rate of Y chromatin was studied along with the morphological change of the tissue. . By the HE staining observation about the dental pulp, left at the room temperature, it was revealed that the pulp tissue can be well preserved for a long time because of natural drying. In both cases the degree of staining declined with the lapse of time after the removal of tooth. The author concluded that the appearance rate of Y chromatin is more in the male pulp tissue when stained immediately after extraction.6
3.Das.N,Gorea RK, Gargi J, Singh JR (2004) conducted a study on pulp of 90 teeth from cadavers extracted at various times after death. They prepared two separate smears from pulp tissue of each patient which were stained with Harris Haematoxylin & Eosin and a fluorescent dye named Quinacrine dihydrochloride respectively. They concluded thatthe Barr body is present in females and F body is present in males.7
4.VeeraraghavanG, Lingappa A, ShankaraSP, GowdaPM, Bastian TSAhmed M (2009) conducted a study on pulp tissue of 60 maxillary and mandibular premolars and permanent molar teeth. The cells stained with Quinacrinedihydrochloride were observed with fluorescent microscope for fluorescent body. Gender was determined by identification of Y chromosome fluorescence in dental pulp. They concluded that the fluorescent Y body test is a simple and cost effective technique for gender identification.1
5.Suazo GI, Roa H I, Cantin L M. (2010) conducted a study using the pulp tissue of 40 premolar and molar teeth. Theyprocessed the pulp tissues, stained withHaematoxylin and Eosin staining and concluded that the Barr body is seen in the pulp tissue of females.8
6.3. Objectives of the study
1. To evaluate the positivity of fluorescent body in the cells of pulp tissue.
2. To evaluate the reliability of sex determination from pulp tissueusing fluorescent body
test.
7. Materials and methods:
7.1 Source of data:
Extracted teeth collected from department of Oral and Maxillofacial Surgery, AECS Maaruti College of Dental Sciences and Research Centre. The details of patients from whom teeth have been extracted will be collected from the same department.
7.2 Method of collection of data
7.2.1 Sample Size:
60 teeth(maxillary and mandibular premolars; 30 from male patients and 30 from female patients) extracted for orthodontic treatment purpose will be selected for study purpose.
7.2.2Study Material:
Extracted teeth,20% Acetic acid,Absolute methanol, Quinacrinedihydrochloride dye, double distilled water, Mcllvaine’s buffer, Magnesium chloride, glass slides, glycerol, fluorescent microscope.
7.2.3Study Method:
The study will be done on 60 maxillary and mandibular premolar teeth free of dental caries, but indicatedfor orthodontic extraction. The pulp will be removed with a sterile curette and kept dry in a sterile test tube.
Staining of dental pulp
The pulp tissue will be transferred into small mortar with a needle and 0.5 ml of 20% acetic acid will be used to soften the dental pulp, which will then be crushed to separate the cells. The cells will be collected on a glass slide and stained with Quinacrine dihydrochloride.
Then a drop of glycerol will be added and cover slip will be placed. The slide will be given to two independent observers for observation of fluorescent body under fluorescent microscope to reduce inter-observer bias. The fluorescent body appears in the nucleus of male cells.
7.2.4Study Design
Single blind study
7.3Inclusion criteria:
1. Premolar teeth
2. Sound and non decayed teeth
7.4 Exclusion criteria:
1. Teeth with extensive decay involving pulp.
2. Teeth with periapical pathology
3. Endodontically treated teeth
7.5 Does the study require any investigation or interventions to be conducted on patients or other humans or animals? If so, please describe briefly.
Yes
7.6 Has ethical clearance been obtained from your institution for this study?
Yes.
8List of references:
1)Veeraraghavan G, Lingappa A, Shankara SP, Gowda PM, Bastian TS,Ahmed MDetermination of sex from tooth pulp tissue Libyan J Med 2010; 5: 5084 - DOI:10.3402/ljm.v5i0.5084
2)Ramenzoni LL, Line SRP. Automated biometrics-based personal identification of the Hunter Schreger bands of dental enamel. Proc Biol Sci. 2006; 273: p.1155-8.
3)Shetty.M, Premalatha KOriginal research paperABO Blood Grouping from Tooth Material. J Indian Acad Forensic Med, 2010; 32(4): p.336-9
4)Malaver PC, Yunis JJ. Different dental tissues as source of DNA for human identification in forensic cases. Croat Med J. 2003; 44: p.306-9.
5)Pillay VV. MKR Krishnan’s hand book of forensic medicine and toxicology. 12th ed. Hyderabad, India: Paras Publications; 2001: p. 34.
6)Adachi H. (1) Studies on sex determination using human dental pulp; (2) Sex determination of teeth left in a room. Nippon Hoigaku Zasshi. 1989; 43: p.27-39.
7)Das.N,Gorea RK, Gargi J, Singh.J.R, Sex determination from pulpal tissue.J Indian Acad Forensic Med, 2004; 26(2): p.50-54
8)Suazo, G. I, Roa HI& Cantín LM. Sex Chromatin in Dental Pulp. Performance of Diagnosis Test and Gold Standard Generation Int.J.Morphol., 2010; 28(4):p.1093-1096.
9)Bailoor DN, Nagesh KS. Fundamentals of oral medicine and radiology. 2nd ed. New Delhi, India: Jaypee Publishers 2001; p. 333.
10)Yamada Y,Yamamoto K,Yoshii T,Ishiyama I. Analysis of DNA from tooth and application to forensic dental medicineNihon Hoigaku Zasshi.1989; Oct; 43(5): p.420-3.
9. / Signature of Candidate10. / Remarks of the guide :
11. / 11.1Name & Designation of
Guide : / Dr. PUSHPALATHA MAHESH
PROFESSOR,
Department of Oral and Maxillofacial Pathology,
AECS Maaruti College of Dental Sciences and
Research Centre,
Bangalore – 560076
11.2 Signature :
11.3 Co-Guide
11.4 Signature :
11.5 Head of Department / Dr. P.SHARADA
PROFESSOR & HEAD,
Department of Oral and Maxillofacial Pathology,
AECS Maaruti College of Dental Sciences and
Research Centre,
Bangalore – 560076
11.6 Signature :
12. / 12.1 Remarks of the Chairman & Principal :
12.2 Signature
PATIENT CONSENT FORM
Name:
Age:
Sex:
Address:
Tooth extracted:
I hereby give my consent to the Department of Oral and Maxillofacial Pathology to use the pulp tissue from my extracted tooth for research purpose.
Signature of the patient
Date
RAJIV GANDHI UNIVERSITY OF
HEALTH SCIENCES
BANGALORE, KARNATAKA
MASTER OF DENTAL SURGERY (MDS)
ORAL AND MAXILLOFACIAL PATHOLOGY
2013
AECS MAARUTI COLLEGE OF
DENTAL SCIENCES
AND RESEARCH CENTRE, BANGALORE --76