Validation of a High-Throughput Dicentric Chromosome Assay Using Complex Radiation Exposures.

Validation of biodosimetry assays is routinely performed using primarily orthovoltage irradiators at a conventional dose rate of approximately 1 Gy/min. However, incidental/ accidental exposures caused by nuclear weapons can be more complex. The aim of this work was to simulate the DNA damage effects mimicking those caused by the detonation of a several kilotons improvised nuclear device (IND). For this, we modeled complex exposures to: 1. a mixed (photons + IND-neutrons) field and 2. different dose rates that may come from the blast, nuclear fallout, or ground deposition of radionuclides (ground shine). Additionally, we assessed whether myeloid cytokines affect the precision of radiation dose estimation by modulating the frequency of dicentric chromosomes. To mimic different exposure scenarios, several irradiation systems were used. In a mixed field study, human blood samples were exposed to a photon field enriched with neutrons (ranging from 10% to 37%) from a source that mimics Hiroshima's A-bomb's energy spectrum (0.2-9 MeV). Using statistical analysis, we assessed whether photons and neutrons act in an additive or synergistic way to form dicentrics. For the dose rates study, human blood was exposed to photons or electrons at dose rates ranging from low (where the dose was spread over 32 h) to extremely high (where the dose was delivered in a fraction of a microsecond). Potential effects of cytokine treatment on biodosimetry dose predictions were analyzed in irradiated blood subjected to Neupogen or Neulasta for 24 or 48 h at the concentration recommended to forestall manifestation of an acute radiation syndrome in bomb survivors. All measurements were performed using a robotic station, the Rapid Automated Biodosimetry Tool II, programmed to culture lymphocytes and score dicentrics in multiwell plates (the RABiT-II DCA). In agreement with classical concepts of radiation biology, the RABiT-II DCA calibration curves suggested that the frequency of dicentrics depends on the type of radiation and is modulated by changes in the dose rate. The resulting dose-response curves suggested an intermediate dicentric yields and additive effects of photons and IND-neutrons in the mixed field. At ultra-high dose rate (600 Gy/s), affected lymphocytes exhibited significantly fewer dicentrics (P < 0.004, t test). In contrast, we did not find the dose-response modification effects of radiomitigators on the yields of dicentrics (Bonferroni corrected P > 0.006, ANOVA test). This result suggests no bias in the dose predictions should be expected after emergency cytokine treatment initiated up to 48 h prior to blood collection for dicentric analysis.

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