Gamma-H2AX biodosimetry for use in large scale radiation incidents: comparison of a rapid ‘96 well lyse/fix’ protocol with a routine method
Following a radiation incident, preliminary dose estimates made by γ-H2AX foci analysis can supplement the early triage of casualties based on clinical symptoms. Sample processing time is important when many individuals need to be rapidly assessed. A protocol was therefore developed for high sample...
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PeerJ Inc.
2014-03-01
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author | Jayne Moquet Stephen Barnard Kai Rothkamm |
author_facet | Jayne Moquet Stephen Barnard Kai Rothkamm |
author_sort | Jayne Moquet |
collection | DOAJ |
description | Following a radiation incident, preliminary dose estimates made by γ-H2AX foci analysis can supplement the early triage of casualties based on clinical symptoms. Sample processing time is important when many individuals need to be rapidly assessed. A protocol was therefore developed for high sample throughput that requires less than 0.1 ml blood, thus potentially enabling finger prick sampling. The technique combines red blood cell lysis and leukocyte fixation in one step on a 96 well plate, in contrast to the routine protocol, where lymphocytes in larger blood volumes are typically separated by Ficoll density gradient centrifugation with subsequent washing and fixation steps. The rapid ‘96 well lyse/fix’ method reduced the estimated sample processing time for 96 samples to about 4 h compared to 15 h using the routine protocol. However, scoring 20 cells in 96 samples prepared by the rapid protocol took longer than for the routine method (3.1 versus 1.5 h at zero dose; 7.0 versus 6.1 h for irradiated samples). Similar foci yields were scored for both protocols and consistent dose estimates were obtained for samples exposed to 0, 0.2, 0.6, 1.1, 1.2, 2.1 and 4.3 Gy of 250 kVp X-rays at 0.5 Gy/min and incubated for 2 h. Linear regression coefficients were 0.87 ± 0.06 (R2 = 97.6%) and 0.85 ± 0.05 (R2 = 98.3%) for estimated versus actual doses for the routine and lyse/fix method, respectively. The lyse/fix protocol can therefore facilitate high throughput processing for γ-H2AX biodosimetry for use in large scale radiation incidents, at the cost of somewhat longer foci scoring times. |
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language | English |
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spelling | doaj.art-2b5ba7be6d47423fadfce0f9f74a7a6e2023-12-03T10:04:10ZengPeerJ Inc.PeerJ2167-83592014-03-012e28210.7717/peerj.282282Gamma-H2AX biodosimetry for use in large scale radiation incidents: comparison of a rapid ‘96 well lyse/fix’ protocol with a routine methodJayne Moquet0Stephen Barnard1Kai Rothkamm2Public Health England, Centre for Radiation Chemical and Environmental Hazards, Chilton, Didcot, Oxfordshire, UKPublic Health England, Centre for Radiation Chemical and Environmental Hazards, Chilton, Didcot, Oxfordshire, UKPublic Health England, Centre for Radiation Chemical and Environmental Hazards, Chilton, Didcot, Oxfordshire, UKFollowing a radiation incident, preliminary dose estimates made by γ-H2AX foci analysis can supplement the early triage of casualties based on clinical symptoms. Sample processing time is important when many individuals need to be rapidly assessed. A protocol was therefore developed for high sample throughput that requires less than 0.1 ml blood, thus potentially enabling finger prick sampling. The technique combines red blood cell lysis and leukocyte fixation in one step on a 96 well plate, in contrast to the routine protocol, where lymphocytes in larger blood volumes are typically separated by Ficoll density gradient centrifugation with subsequent washing and fixation steps. The rapid ‘96 well lyse/fix’ method reduced the estimated sample processing time for 96 samples to about 4 h compared to 15 h using the routine protocol. However, scoring 20 cells in 96 samples prepared by the rapid protocol took longer than for the routine method (3.1 versus 1.5 h at zero dose; 7.0 versus 6.1 h for irradiated samples). Similar foci yields were scored for both protocols and consistent dose estimates were obtained for samples exposed to 0, 0.2, 0.6, 1.1, 1.2, 2.1 and 4.3 Gy of 250 kVp X-rays at 0.5 Gy/min and incubated for 2 h. Linear regression coefficients were 0.87 ± 0.06 (R2 = 97.6%) and 0.85 ± 0.05 (R2 = 98.3%) for estimated versus actual doses for the routine and lyse/fix method, respectively. The lyse/fix protocol can therefore facilitate high throughput processing for γ-H2AX biodosimetry for use in large scale radiation incidents, at the cost of somewhat longer foci scoring times.https://peerj.com/articles/282.pdfGamma-H2AX fociDNA double-strand breakBiological dosimetryIonising radiationTriageBlood sample processing |
spellingShingle | Jayne Moquet Stephen Barnard Kai Rothkamm Gamma-H2AX biodosimetry for use in large scale radiation incidents: comparison of a rapid ‘96 well lyse/fix’ protocol with a routine method PeerJ Gamma-H2AX foci DNA double-strand break Biological dosimetry Ionising radiation Triage Blood sample processing |
title | Gamma-H2AX biodosimetry for use in large scale radiation incidents: comparison of a rapid ‘96 well lyse/fix’ protocol with a routine method |
title_full | Gamma-H2AX biodosimetry for use in large scale radiation incidents: comparison of a rapid ‘96 well lyse/fix’ protocol with a routine method |
title_fullStr | Gamma-H2AX biodosimetry for use in large scale radiation incidents: comparison of a rapid ‘96 well lyse/fix’ protocol with a routine method |
title_full_unstemmed | Gamma-H2AX biodosimetry for use in large scale radiation incidents: comparison of a rapid ‘96 well lyse/fix’ protocol with a routine method |
title_short | Gamma-H2AX biodosimetry for use in large scale radiation incidents: comparison of a rapid ‘96 well lyse/fix’ protocol with a routine method |
title_sort | gamma h2ax biodosimetry for use in large scale radiation incidents comparison of a rapid 96 well lyse fix protocol with a routine method |
topic | Gamma-H2AX foci DNA double-strand break Biological dosimetry Ionising radiation Triage Blood sample processing |
url | https://peerj.com/articles/282.pdf |
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