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Public Security S&T Symposium 2009 CRTI 06-0186RD Novel DNA-Based Radiological Dosimetry Technology Presented by: Isabelle Charlebois, CRI June 17th, 2009.

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Presentation on theme: "Public Security S&T Symposium 2009 CRTI 06-0186RD Novel DNA-Based Radiological Dosimetry Technology Presented by: Isabelle Charlebois, CRI June 17th, 2009."— Presentation transcript:

1 Public Security S&T Symposium 2009 CRTI RD Novel DNA-Based Radiological Dosimetry Technology Presented by: Isabelle Charlebois, CRI June 17th, 2009

2 Introduction and Objectives The purpose of this project is to develop a personal and wearable dosimeter using a highly-innovative approach based on the specific recognition of DNA damage with a bio-sensitive polymer hybrid This three year project follows up on CRTI RD and CRTI RD The proposed research will seek to develop a two component fluidic device comprising: –1) a wearable sensor module consisting of a solid support harboring covalently bound known amounts of target nucleic acid of predefined length and having a specific terminal sequence tag –2) a portable detector module in which the wearable module will be inserted for detection of the cleaved nucleic acid by measuring the fluorescence emitted by the polymer-based sensor ST IC 2

3 Introduction - DNA Dosimeter Sensor Module Surface Anchor 70-base oligonucleotide 20-base Tag Sequence Radiation Break Released broken DNA ST IC 3

4 FCR aggregates (nanometers) Polymer DNA probe Superlighting (nanometers) Released broken DNA Introduction - DNA Dosimeter Detector Module 4

5 FCR aggregates (nanometers) FCR aggregates (nanometers) Microbeads (micrometers) Released broken DNA Superlighting on beads ST IC 5

6 Introduction - DNA Dosimeter Unique Features Detection of DNA damages in a synthetic system –Compatible with all radiation types affecting human DNA –No constraint or variables from other biological mechanisms such as DNA repair A novel detection technology –Discovered and developed under previous CRTI grants –Unique and sensitive signal amplification method This DNA dosimeter will help Canadian authorities in assessing health risks when faced with radiological or nuclear event ST IC 6

7 Released ssDNA Surface Tag 20 bases Real-time PCR Analysis PCR-20 for tag detection, based on SybrGreen Surface Tag PCR-70 for 70 bases oligonucleotide detection, based on a Taqman probe Sybr Green fluorescence dsDNA only fluorophore quencher Real-time PCR Analysis Fluorophore released 70 bases Results - DNA Dosimeter Development of 2 real-time PCR assays ST IC 7

8 Results - DNA Dosimeter Radiation Sensitivity of the DNA Target Gamma-irradiation of unbound DNA in solution At 10 Gy, 35% of the DNA is not suitable for PCR amplification ST IC 8

9 Results - DNA Dosimeter Sensor module – Background DNA Not all DNA stays on the surface (irradiated or not) First generation of sensor module = 0.01% of full-length DNA recovered (without irradiation) 0.01% of copies = copies Surface Bound DNA Released broken DNA + Background DNA ST IC 9

10 Results - DNA Dosimeter Sensor Module – Background DNA Not all DNA stays on the surface (irradiated or not) First generation of sensor module = 0.01% of full-length DNA recovered (without irradiation) Improved procedures (chemical bounding, washing treatments, other type of exposure device) reduced background DNA up to % LinkDNA surfaceBackgroundRatio (background/bounded DNA) EDC-NHS slides2 x 10 14* copies2 x 10 9 copies1/ % Synthesis columns1 x copies5 x 10 9 copies1/ % *DNA introduced at the beginning, not all DNA is bound to the surface, actual numbers are unknown ST IC 10

11 Results - DNA Dosimeter Irradiation Experiments at CERN Mixed-field irradiation ( 5 Gy) at CERN facilities Increased Tag detection indicates DNA released upon irradiation ST IC 11

12 Results - DNA Dosimeter Detector Module Design & fabrication of microfluidic detection module (beads manipulation, magnetic mixing & high performance capture unit) 1st prototype of microfluidic platform ready (manifold, electro-valves, software development) ST IC 12

13 Results – Detection Platform Prototype for sensitive detection of fluorophore-grafted beads COPLs detection platform: stand-alone (moves the optical head with respect to sample, images detection area, collects images/video files, detects fluorescence) Optical characterization of plastic materials that will be used for the microfluidic readout devices Development of novel fluorescence-based detection strategies for the total analysis of microvolumes at the single molecule level ST IC 13

14 Conclusions - DNA dosimeter Sensor Module: –Proof-of-concept –Tools to quantify DNA –DNA background requires improvement Detector Module: –Microfluidic chips developed to confine fluorophore-grafted beads –Detection platform is ready ST IC 14

15 Next Steps - DNA Dosimeter Sensor Module: –Validate improved exposure module for radiation detection Mixed-field radiation Specific radiation (gamma, neutrons) –Develop post-irradiation treatments to reduce contribution from DNA background Detector Module: –Design a simple fluidic interface to connect microfluidic detection chips with detection platform –Develop / optimize detection strategies for fluorophore-grafted beads confined into microfluidic particle traps –Validate AOB detection in microflluidic chips on the detection platform –Integrate other modules with the detection platform Evaluate the radiation dose range covered by the detection approach and determine its applicability to targeted irradiation fields ST IC 15

16 CRTI RD – DNA dosimeter A multidisciplinary team! Dr. Michel G. Bergeron Centre de recherche en infectiologie (CRI) CHUQ Research Center Molecular Biology Teodor Veres Industrial Materials Institute (IMI) National Research Council Canada Materials / Nanofabrication / Microfluidic Mario Leclerc Research Centre on Advanced Materials (CERMA) Laval University Advanced Materials / Photoactive Polymers Denis Boudreau Centre on Optics, Photonics and Lasers (COPL) Laval University Optics / Photonics / Analytical Chemistry Brent Lewis / Les G. I. Bennett Royal Military College (RMC) Physical Dosimetry / Irradiations Diana Wilkinson Defense R&D Canada-Ottawa (DRDC) Biological Dosimetry Martin Pierre Director General Nuclear Safety (DGNS) Irradiations ST IC 16

17 Questions? Merci Thank you ST IC 17


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