-Dr. Guy Garty Awarded NIAID Contract
Dr. Garty has been awarded a 1.6M$ 3-year contract from NIAID for advanced development of an automated Dicentric Chromosome Assay (DCA).
The manual, lab-based DCA is the gold standard in radiation biodosimetry and is recommended by ISO, IAEA and the FDA. The goal of the NIAID contract is to implement and validate the assay on a commercial high throughput cellular screening platform, increasing throughput by two orders of magnitude compared to the lab implementation, making this important assay applicable to mass triage following a radiological event.
Automating the Dicentric Chromosome Assay (DCA)
Following a large scale radiological or nuclear event, large numbers of people may be exposed to ionizing radiation/s and require subsequent dose-dependent medical management. The Dicentric Chromosome Assay (DCA) is currently the gold standard for clinical radiation biodosimetry: It is recommended for use by the International Atomic Energy Agency (IAEA) and by the International Standards Organization (ISO). The DCA is also recognized by the Food and Drug Administration (FDA) as a reference method for validating other radiation biodosimetry assays.
Dicentric chromosome formation is the result of induced breaks in two chromosomes which, after mis-rejoining, result in a single chromosome entity with two intact centromeres – the dicentric. Because ionizing radiation is a very efficient inducer of DNA double-strand breaks (DSBs) and dicentric formation generally requires induction of DSBs in two chromosomes, dicentrics are a highly specific marker for radiation exposure, particularly in the context of the radiation doses (³1 Gy) relevant to medical management.
The DCA has been successfully employed for radiation biodosimetry in many radiation accidents where the number of people assessed was small. However, as implemented in a standard cytogenetic laboratory, the DCA is too labor intensive to be widely applied when the need is to assess large numbers of individuals. Even large cytogenetic laboratory networks can only analyze some hundreds of samples per day.
Following up on the CRR’s work in development of the RABiT and RABiT-II approaches, Dr. Garty has been awarded a 1.6M$ 3 year contract from NIAID to implement and validate the DCA on our RABiT II system, based on a commercial Perkin Elmer cell::explorer.
High Throughput Screening systems, such as the cell::explorer, use robotics, liquid handling devices and automated imaging to rapidly perform chemical, genetic, or pharmacological tests. The number of commercial HTS machines in universities and industry is rapidly growing as these systems become more affordable and more multi-functional.
As well as their increasing availability, a second major advantage of using commercial HTS systems over both manual systems and also custom-built automated systems is improved quality control. In contrast to dedicated biodosimetry-specific devices which will potentially be unused for long time periods, HTS devices are typically in regular use and undergo regular QA/AC testing and maintenance. These HTS systems also have a broad base of trained users and maintenance personnel ensuring successful operation in time of crisis.
Central to the automation of the DCA is development of an automated scoring algorithm. The "classic" dicentric assay in which metaphase cells are first identified, after which dicentric chromosomes are identified by their morphometric shape is not amenable to high-speed automated image analysis. Our approach is based on fluorescent in situ hybridization (FISH) of the chromosomes. Dicentrics are thus identified as chromosomes containing two green spots (centromeres). This approach is much easier to automate.