Quantification of histone H2AX phosphorylation in white blood cells induced by ex vivo gamma irradiation of whole blood by both flow cytometry and foci counting as a dose estimation in rapid triage
Issued Date
2022-03-01
Resource Type
eISSN
19326203
Scopus ID
2-s2.0-85126857869
Pubmed ID
35320288
Journal Title
PLoS ONE
Volume
17
Issue
3 March
Rights Holder(s)
SCOPUS
Bibliographic Citation
PLoS ONE Vol.17 No.3 March (2022)
Suggested Citation
Wanotayan R., Wongsanit S., Boonsirichai K., Sukapirom K., Buppaungkul S., Charoenphun P., Songprakhon P., Jangpatarapongsa K., Uttayarat P. Quantification of histone H2AX phosphorylation in white blood cells induced by ex vivo gamma irradiation of whole blood by both flow cytometry and foci counting as a dose estimation in rapid triage. PLoS ONE Vol.17 No.3 March (2022). doi:10.1371/journal.pone.0265643 Retrieved from: https://repository.li.mahidol.ac.th/handle/20.500.14594/87561
Title
Quantification of histone H2AX phosphorylation in white blood cells induced by ex vivo gamma irradiation of whole blood by both flow cytometry and foci counting as a dose estimation in rapid triage
Other Contributor(s)
Abstract
A quick, reliable, and reproducible biological assay to distinguish individuals with possible life-threatening risk following radiological or nuclear incidents remains a quest in biodosimetry. In this paper, we examined the use of a γ-H2AX assay as an early dose estimation for rapid triage based on both flow cytometry and image analyses. In the experiment, whole blood from 11 donors was irradiated ex vivo inside a water phantom by gamma rays from Co-60 at 0.51 Gy/min. After the lysis of red blood cells, the white blood cells were collected for immunofluorescence labeling of γ-H2AX, CD45, and nuclear stained for signal collection and visualization. Analysis by flow cytometry showed that the relative γ-H2AX intensities of lymphocytes and granulocytes increased linearly with absorbed doses from 0 to 6 Gy with a large variation among individuals observed above 2 Gy. The relative γ-H2AX intensities of lymphocytes assessed by two different laboratories were highly correlated (ICC = 0.979). Using confocal microscopic images, γH2AX foci were observed to be discretely distributed inside the nuclei and to increase proportionally with doses from 0 to 2 Gy, whereas large plagues of merged foci appeared at 4 and 6 Gy, resulting in the saturation of foci counts above 4 Gy. The number of total foci per cell as well as the number of foci per plane were significantly different at 0 vs 1 and 2 vs 4 Gy doses (p < 0.01). Blind tests at 0.5 Gy and 1 Gy doses showed that dose estimation by flow cytometry had a mean absolute difference of less than 0.5 Gy from the actual value. In conclusion, while flow cytometry can provide a dose estimation with an uncertainty of 0.5 Gy at doses ≤ 1 Gy, foci counting can identify merged foci that are prominent at doses ≥ 4 Gy.