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RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES, BANGALORE, KARNATAKA ANNEXURE II PROFORMA FOR REGISTRATION OF SUBJECT FOR DISSERTATION 1. NAME OF THE CANDIDATE AND ADDRESS: DR.CHAITANYA. R. UPPIN DEPARTMENT OF PEDIATRIC AND PREVENTIVE DENTISTRY, P.M.N.M. DENTAL COLLEGE AND HOSPITAL, BAGALKOT-587101 KARNATAKA. 2.NAME OF THE INSTITUTION: P.M.N.M. DENTAL COLLEGE AND HOSPITAL, BAGALKOT-587101 KARNATAKA 3.COURSE OF STUDY AND SUBJECT: 4. DATE OF ADMISSION: MASTER OF DENTAL SURGERY (M.D.S.) IN PAEDODONTIA 31stMAY 2011 5.TITLE OF THE TOPIC: “DETERMINATION OF ABO BLOOD GROUPS FROM DENTAL PULP AND DENTIN IN DECIDUOUS TEETH” 6. BRIEF RESUME OF THE INTENDED WORK: 6.1 NEED FOR THE STUDY Teeth are invaluable source of personal identification in the field of forensic medicine as they are resistant to environmental assaults. Teeth can survive for long time even after soft and skeletal tissues have been destroyed. Blood grouping from teeth could be a source of personal identification. The use of blood group substance in medico-legal examination is based on the fact that once a group is established in an individual it remains unchanged throughout his/her life1. It was thought to be of interest to apply the ABO blood grouping method to a calcified tissue such as dentin of teeth because it has higher ratio of cell substances than bone2. Pulp tissue is protected from all sides from dental hard tissue and also it has rich vascular supply, blood group antigens are bound to be present in tooth pulp and therefore could be readily available for examination. Most of the studies have used permanent tooth pulp for blood group determination1,7 but only few studies have been undertaken to determine efficacy and reliability of blood grouping using pulp of deciduous dentition. Hence the present study is taken up to determine blood group antigens from pulp and dentin of deciduous tooth, which can be a valuable source for personal identification, when permanent teeth are not available. 6.2 REVIEW OF LITERATURE: A study was done to determine ABO blood grouping on the pulp, dentin and enamel on 35 teeth using absorption elution technique. Twenty teeth were examined within six weeks of extraction and 15 teeth after 6-10 months. It was found that blood grouping on pulp gives fairly good results, whereas the possibilities for correct blood grouping seem to be limited for dentin and debatable for enamel.3 In another study ABO blood group determination was done from dental pulp and saliva for its use in Forensic Odontology. Hundred cases were studied using modified Absorption Elution technique, 93 cases were positive for dental pulp and 79 were positive for saliva.4 Thermo stability of ABO blood group antigens in human dental pulp was described in a study conducted in the year 1978. The ability of the enamel and dentin to protect the ABO antigens from thermal damage was estimated. The calculations shows that dentin and enamel are poor insulators and give inadequate thermal protection to the pulp when external temperature rises to 2000 c and more.5 A similar study was conducted at Mangalore for ABO blood group determination from dental pulp and dentin from sixty teeth taken from dead bodies after 180 days, by using Absorption Elution technique they were able to successfully determine the ABO blood groups only from dental pulp but failed to determine the same from dentin.6 A comparative study was done to correlate the blood group determination by dental pulp and dentin with that of socket blood collected from the same person. The ABO blood group determined by dental pulp correlated well whereas the blood groups from dentin failed to correlate.7 Another study conducted on 183 permanent and 77 deciduous extracted teeth examined after 1-2 months by Elution technique proved that minute amount (6-10 mg) of pulverized tooth material would be sufficient to determine the ABO blood groups.8 6.3 OBJECTIVES OF THE STUDY: 1. To determine ABO blood group from dental pulp and dentin of deciduous teeth after storing the teeth for 6 months. 2. To determine ABO blood group from the socket blood. 3. Correlation of ABO blood group determined by dental pulp, dentin and socket blood. 7. MATERIALS AND METHODS: 7.1 SOURCE OF THE DATA 60 children between age group 3-12 yrs coming to the Department of Pedodontics and Preventive Dentistry, P. M. N. M. Dental College and Hospital, Bagalkot for extraction of over retained teeth or extraction of deciduous teeth for orthodontic treatment are randomly chosen for this study. INCLUSION CRITERIA: 1. Over retained teeth. 2. Deciduous Teeth indicated for orthodontic extraction. EXCLUSION CRITERIA: 1. Carious teeth.7 7.2 METHODS OF COLLECTING DATA 60 children are selected randomly coming to the Department of Pedodontics and Preventive Dentistry, P. M. N. M. Dental College and Hospital, Bagalkot for extraction of over retained teeth or extraction of deciduous teeth for orthodontic treatment. MATERIALS: 1. Mouth mirror & Dental probe. 2. Anticoagulant solution EDTA 3. Fresh normal saline. 4. Antisera A and B. 5. Gauze pieces, numbered bottles. 6. Lathe, carborundum disc, spoon excavator. 7. Straight fissure bur. 8. Centrifuge, Hot water bath, Incubator. 9. A and B Red Blood cell suspension. 10. Microscope METHOD: The study group shall comprise of 60 patients requiring extraction for overretained teeth or deciduous teeth for orthodontic reasons. A brief case history with relevant medical history will be recorded and a detailed clinical examination under artificial illumination will be done. Extraction procedure is carried under local anesthesia following an aseptic precaution. After extraction, the socket is compressed with a sterile gauze piece and socket blood will be collected for ABO blood group determination which serves as a control group for the study. Anticoagulant is added to the socket blood and one drop of this blood is placed on the slide and a drop of Antiserum-A is added, if agglutination occurs on addition of Antiserum-A, blood group shall be recorded as A and vice versa. The extracted teeth will be washed under running water and debris shall be removed with the probe and wiped with gauze and kept in bottles which are numbered for identification these teeth are stored for a span of 6 months7. The blood grouping is performed by Absorption Elution procedure using dental pulp and dentin and later compared with control sample7. The tooth is completely trimmed to remove the enamel and cementum with lathe. The tooth is further split vertically with carborundum disc and the dental pulp is scooped out with spoon excavator, the remaining tooth consisting of dentin, shall be pulverized with straight fissure bur. The pulverized tooth powder is put in two test tubes, to each of these test tubes three drops of antiserum A and B shall be added and ensuring that the test samples are being sufficiently soaked with antiserum for two and half hours and left standing at room temperature. After removing antiserum, each sample shall be washed three times with cold saline solution and centrifuged and the supernatant is sucked with pipette. Then two drops of fresh saline is added to the sample and the test tube is heated in a water bath at 50-55°c for 10 minutes to elute the antibodies. A drop of 0.5% A or B group RBC suspension is immediately put into each respective test tube of known blood and the samples shall be put in humified recipient and shall be incubated at 37°c for 30min to enhance agglutination and after this procedure, it shall be centrifuged at 1,500- 2000rpm for 1 min. By gentle shaking of the test tube the presence or absence of RBC agglutination shall be ascertained with microscope at magnification of 100X. Actual blood groups determined from the socket blood, dental pulp and dentin shall be correlated using appropriate statistical methods. 7.3 DOES THE STUDY REQUIRE ANY INVESTIGATION OR INTERVENTION TO BE CONDUCTED ON PATIENTS OR OTHER HUMANS OR ANIMALS? IF SO PLEASE BRIEFLY SPECIFY. Yes. 7.4 HAS ETHICAL CLEARANCE BEEN OBTAINED FROM YOUR INSTITUTION? Yes, a copy is attached. 8. LIST OF REFERANCES: 1. Neiders ME. Blood group determinations in forensic dentistry. Dent Clin North Am.1977; 21(1):99-111 2. Kramer Ivor RH. An examination of Dentin for A and B Bloodgroup antigens by mixed agglutination technique. Proceedingsof Royal society of medicine. 1957; 50:677-78 3. Smeets B. ABO blood grouping on tooth material. Forensic Sci Int. 1991; 50(2):277-84. 4. Parekh, K Sansare, AG Malwankar, PG Gore. ABO blood group determination from dental pulp and saliva for its use in forensic Odontology. Journal of Indian Academy of Oral Medicine and Radiology 1994; 2:17-20. 5. Anna-Krystyna Karszun. Thermo stability of ABH (O) blood antigens in human teeth. Journal Forensic Science 1978; 11: 231-39. 6. Dr. Mahabaleshshetty, Dr. Premlatha K. ABO blood grouping from tooth material. Journal of Indian forensic medicine 2010; 32(4):336-338. 7. SushminiBallal, Maria Priscilla David. Determination of ABO blood grouping from dentin and pulp. Pakistan Oral and dental journal 2011; 31( 1):3-6. 8. Hideo Takata. Blood group identification from hard dental tissues by elution test. International Journal of Forensic dentistry1974; 2(4):43-47.