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Abstract: 141-1

141-1

PROTOCOL OF COMBINED CYTOGENETIC TECHNIQUES FOR TRIAGE AND DOSE ESTIMATION IN LARGE-SCALE NUCLEAR AND RADIOLOGICAL ACCIDENTS

Authors:
Caio Ferreira de Oliveira (UFPE - Universidade Federal de Pernambuco, CRCN/NE - Centro Regional de Ciências Nucleares do Nordeste) ; Gael Freires de Lima (UFPE - Universidade Federal de Pernambuco, CRCN/NE - Centro Regional de Ciências Nucleares do Nordeste) ; Suy Ferreira Hwang (CRCN/NE - Centro Regional de Ciências Nucleares do Nordeste) ; Amanda Silva de França (CRCN/NE - Centro Regional de Ciências Nucleares do Nordeste) ; Fernanda Celina Tavares de Moraes (UFPE - Universidade Federal de Pernambuco, CRCN/NE - Centro Regional de Ciências Nucleares do Nordeste) ; Akiria Ohana Torreão (UFPE - Universidade Federal de Pernambuco) ; Fabiana Farias de Lima (CRCN/NE - Centro Regional de Ciências Nucleares do Nordeste)

Abstract:

Ionizing Radiation (IR) is used in assorted social sectors due to its different aspects, whether through diagnostic evaluation and treatments, or by industrial processes. Despite this, inadequate exposure of IR leads to even large-scale radiological and nuclear accidents that cause DNA damage when in contact with the human organism[1]. Thus, Biological Dosimetry uses cytogenetic techniques, such as, for example, Dicentric Technique (Gold Standard) and Cytokinesis Block Micronucleus Assay (CBMN)[2], which are used as tools to estimate absorbed doses and in triage scoring in a mass casualty event through the cultivation of circulating lymphocytes of peripheral blood. However, these methods are time-consuming both in cell cultivation and in microscopic analysis and consequently there is a delay in the necessary laboratory response so the medical group can give the specific care and treatment. In this sense, there is an urgent need to create a protocol with the two parallel techniques[3] most used in Biological Dosimetry with the aim of reducing analysis time, laboratory costs, and quickly give a feedback to the medical team by performing dosimetric triage and estimate absorbed dose using the two most commonly techniques in a unique procedure.Therefore,blood samples were collected from a volunteer and irradiation was performed in a 60Co source on the Gammacel 220 irradiator at the Department of Nuclear Energy at UFPE with absorbed doses of 0.5Gy,0.75Gy,2Gy and 3Gy. Three different times of cell culture were analyzed, 48h, 58h and 72h. After irradiation, the culture was started by supplementation with RPMI and phytohemagglutinin to stimulate the cell proliferation and adding Cytochalasin B at 24h to block the Cytokinesis and Colchicine 3h before the end of the respective cell culture times so it can stop the cell division at metaphase after that comes a hypotony step with KCl and fixation and washing with acetic acid and methanol to make the slides that are bleached with 5% Giemsa. After these phases,microscopic analysis confirmed that the combining cytogenetic techniques are effective for triage by demonstrating Metaphases with cell cycle differentiation (M1 and M2) and chromosomal alterations, beyond Binucleated Cells (BN) with Micronuclei.  Cultures at 48h and 72h the  scored a lower number of cells than that recommended by the IAEA (International Atomic Energy Agency) of 1000 metaphases and 1000 BN, as the sample number of cells analyzed showed minor cell proliferation probably due to cultivation time reducing the cell yields. However, in cases of triage in radiological/nuclear accidents, the number of BN drops to 200, since as it is a more sensitive technique, it is easier to obtain a dose estimate. For this, an intermediate time is being tested and the analysis already demonstrates a higher number of BN indicating a rapid response.

Keywords:
 Biodosimetry, Cytogenetics, Dicentric, Micronucleus, Radioprotection