[en] A system for scoring dicentric chromosomes by image analysis comprised fully automatic location of mitotic cells, automatic retrieval, focus and digitisation at high resolution, automatic rejection of nuclei and debris and detection and segmentation of chromosome clusters, automatic centromere location, and subsequent rapid interactive visual review of potential dicentric chromosomes to confirm positives and reject false positives. A calibration set of about 15000 cells was used to establish the quadratic dose response for ⁶⁰Co γ-irradiation. The dose-response function parameters were established by a maximum likelihood technique, and confidence limits in the dose response and in the corresponding inverse curve, of estimated dose for observed dicentric frequency, were established by Monte Carlo techniques. The system was validated in a blind trial by analysing a test comprising a total of about 8000 cells irradiated to 1 of 10 dose levels, and estimating the doses from the observed dicentric frequency. There was a close correspondence between the estimated and true doses. The overall sensitivity of the system in terms of the proportion of the total population of dicentrics present in the cells analysed that were detected by the system was measured to be about 40%. This implies that about 2.5 times more cells must be analysed by machine than by visual analysis. Taking this factor into account, the measured review time and false positive rates imply that analysis by the system of sufficient cells to provide the equivalent of a visual analysis of 500 cells would require about 1 h for operator review. (author). 20 refs.; 4 figs.; 5 tabs

No abstract available

[en] Highlights: • Results of the γ-H2AX assay agree with EWBD better than the DCA results. • The γ-H2AX assay is a valid, precise and rapid alternative to DCA. - Abstract: The γ-H2AX assay was investigated as an alternative to the time-consuming dicentric chromosome assay (DCA). Radiation doses to 25 radiotherapy patients were estimated in parallel by DCA and the γ-H2AX assay. The γ-H2AX assay yielded doses in line with the calculated equivalent whole body doses in 92% of the patients, whereas the success rate of DCA was only 76%. The result shows that the γ-H2AX assay can be effectively used as a rapid and more precise alternative to DCA.

[en] Bio-dosimetry is an essential tool for providing timely assessments of radiation exposure, particularly when physical dosimetry is unavailable or unreliable. For mass-casualty events involving public exposure to ionising radiation, it is paramount to rapidly provide this dose information for medical management of casualties. The dicentric chromosome assay is currently the most reliable accepted method for bio-dosimetry; however, in a mass-casualty scenario, the throughput of this assay will be challenged by its time-consuming nature and the specific expertise required. To address this limitation, many countries have established expertise in cytogenetic bio-dosimetry and started developing surge capabilities through setting up regional networks to deal with emergency situations. To capitalise on this growing expertise and organise it into an internationally coordinated laboratory network, the World Health Organization has created and launched a global bio-dosimetry network (BioDoseNet). In order to determine the existing capacity of BioDoseNet member laboratories, including their expertise and in vivo experience, involvement in national and international activities, problems, needs and prospects, an in-depth survey was conducted. These survey results provide significant information on the current state of emergency cytogenetic bio-dosimetry capabilities around the world. (authors)

[en] The peripheral blood lymphocyte chromosomes of the Filipino, the carabao, the goat, and the ricefield rat, four mammalian species found abundant in the environs of the Philippine Nuclear Power Plant (PNPP-I) in Morong, Bataan, were studied to establish their radiosensitivities and to explore their possible use as biological indicator of radiation effects. The four mammalian species were found to be radiosensitive. Chromosome aberration was induced by gamma-irradiation of whole venous blood. By cytogenetic technique the aberrant chromosomes were evaluated. The aberrant chromosomes may be categorized into chromatid and chromosome types. Of the types seen, it was concluded that dicentrics are the most reliable indicator of radiation effects. In the course of this study a suitable medium was formualted and was found competitive with a commercially prepared medium. The radiosensitivity of the four species based on the frequency of dicentrics differs from each other. A calibration curve was constructed to relate exposure to the observed incidence of dicentrics. This curve is very important in the calculation of the dose corresponding to the observed dicentric yield in case of accidental release of radioactivity in the PNPP-I site. (author)

[en] Dicentric Chromosome Assay (DCA) is considered to be the 'gold standard' for personalized dose assessment in humans after accidental or incidental radiation exposure. Although this technique is superior to other cytogenetic assays in terms of specificity and sensitivity, its potential application to radiation mass casualty scenarios is highly restricted because DCA is time consuming and labor intensive when performed manually. Therefore, it is imperative to develop high throughput automation techniques to make DCA suitable for radiological triage scenarios. At the Cytogenetic Biodosimetry Laboratory in Oak Ridge, efforts are underway to develop high throughput automation of DCA. Current status on development of various automated cytogenetic techniques in meeting the biodosimetry needs of radiological/nuclear incident(s) will be discussed

[en] Methods and procedures for generating, interpreting and scoring the frequency of dicentric chromosomes vary among cytogenetic biodosimetry laboratories (CBLs). This variation adds to the already considerable lack of precision inherent in the dicentric chromosome assay (DCA). Although variability in sample collection, cell preparation, equipment and dicentric frequency scoring can never be eliminated with certainty, it can be substantially minimized, resulting in reduced scatter and improved precision. Use of standard operating procedures and technician exchange may help to mitigate variation. Although the development and adoption of international standards (ISO 21243 and ISO 19238) has helped to reduce variation in standard operating procedures (SOPs), all CBLs must maintain process improvement, and those with challenges may require additional assistance. Sources of variation that may not be readily apparent in the SOPs for sample collection and processing include variability in ambient laboratory conditions, media, serum lot and quantity and the use of particular combinations of cytokines. Variability in maintenance and calibration of metafer equipment, and in scoring criteria, reader proficiency and personal factors may need to be addressed. The calibration curve itself is a source of variation that requires control, using the same known-dose samples among CBLs, measurement of central tendency, and generation of common curves with periodic reassessment to detect drifts in dicentric yield. Finally, the dose estimate should be based on common scoring criteria, using of the z-statistic. Although theoretically possible, it is practically impossible to propagate uncertainty over the entire calibration curve due to the many factors contributing to variance. Periodic re-evaluation of the curve is needed by comparison with newly published curves (using statistical analysis of differences) and determining their potential causes. (author)

[en] The harlequin-staining method has been used to study the cell kinetic of goat peripheral blood lymphocytes stimulated by phytohemagglutinin and to assess their radiosensitivity. At 48 h, the standardized culture time employed for human lymphocytes, 71% of the goat lymphocytes are in first mitosis, 23% are in second mitosis and 5% in third. Irradiation with 200 rads X-rays induces an average of 24,5 dicentric chromosomes per hundred cells in first mitosis

[en] 'Full text:' It is widely accepted that the dicentric chromosome assay (DCA) is currently the most sensitive and radiation-specific assay for measuring biological damage due to exposure to ionizing radiation. However, the utility of this assay following radiological or nuclear accidents is limited due to the time- and expertise-intensive nature of the microscopic scoring. As a strategy to increase throughput for this assay, a new scoring technique (termed DCA QuickScan) has been validated as an alternative rapid scoring approach. Unlike conventional DCA scoring, the individual centromeres are not counted when scoring using DCA QuickScan criteria, rather the metaphase spread is quickly examined (< 10 seconds) to confirm that it appears to be complete. If no damage is obvious during that examination, then the spread is scored as normal. If damage is observed (i.e. fragments, visible rings and/or dicentrics), the scorer carefully enumerates the damage. Each dicentric must be accompanied by an acentric fragment to reduce the chance of mistaking overlapping chromatids with true dicentrics. Evaluating spread morphology is a useful parameter for QuickScan scoring, where round or oval spreads are most reliably found to contain a complete chromosome compliment. As well, the use of an automated slide-maker has proven useful for providing spreads of consistent quality for use in QuickScan scoring. In a previous study, triage-quality conventional DCA and DCA QuickScan analyses were compared based upon scoring a minimum of 50 metaphase cells or 30 dicentrics by 9-15 scorers across 4 laboratories. Results from this pilot study indicated that QuickScan scoring was as accurate, and much faster than, conventional DCA analysis (a 6-fold decrease in scoring time was realized using DCA QuickScan). As a follow-up study, and as validation of this scoring strategy, these scoring methods have been compared by generating full dose response curves with 1000 metaphases or 200 dicentrics scored for each dose point. Blood samples were exposed to 0-4 Gy of gamma radiation, cultured by standard methods and scored using either conventional DCA or DCA QuickScan criteria. The time required for construction of the dose response curves, and the sensitivity of DCA QuickScan scoring in comparison to conventional DCA scoring, will be reported. (author)

[en] Testing and validation of biodosimetry assays is routinely performed using conventional dose rate irradiation platforms, at a dose rate of approximately 1 Gy/min. In contrast, the exposures from an improvised nuclear device will be delivered over a large range of dose rates with a prompt irradiation component, delivered in less than 1 μs, and a protracted component delivered over hours and days. We present preliminary data from a large demographic study we have undertaken for investigation of age, sex and dose rate effects on dicentric and micronucleus yields. Our data demonstrate reduced dicentric and micronucleus yields at very high dose rates. Additionally, we have seen small differences between males and females, with males having slightly fewer micronuclei and slightly more dicentrics than females, at high doses (authors)