Establishment of in vitro Calibration Curve for 60Co-γ-rays Induced Phospho-53BP1 Foci, Rapid Biodosimetry and Initial Triage, and Comparative Evaluations With γH2AX and Cytogenetic Assays
A rapid and reliable method for biodosimetry of populations exposed to ionizing radiation in the event of an incident or accident is crucial for initial triage and medical attention. DNA-double strand breaks (DSBs) are indicative of radiation exposure, and DSB-repair proteins (53BP1, γH2AX, ATM, etc...
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Frontiers Media S.A.
2022-08-01
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| Series: | Frontiers in Public Health |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fpubh.2022.845200/full |
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| author | Rajesh Kumar Chaurasia Rajesh Kumar Chaurasia Kapil B. Shirsath Utkarsha N. Desai Nagesh N. Bhat Nagesh N. Bhat B. K. Sapra B. K. Sapra |
| author_facet | Rajesh Kumar Chaurasia Rajesh Kumar Chaurasia Kapil B. Shirsath Utkarsha N. Desai Nagesh N. Bhat Nagesh N. Bhat B. K. Sapra B. K. Sapra |
| author_sort | Rajesh Kumar Chaurasia |
| collection | DOAJ |
| description | A rapid and reliable method for biodosimetry of populations exposed to ionizing radiation in the event of an incident or accident is crucial for initial triage and medical attention. DNA-double strand breaks (DSBs) are indicative of radiation exposure, and DSB-repair proteins (53BP1, γH2AX, ATM, etc.) are considered sensitive markers of DSB quantification. Phospho-53BP1 and γH2AX immunofluorescence technique serves as a sensitive, reliable, and reproducible tool for the detection and quantification of DSB-repair proteins, which can be used for biological dose estimations. In this study, dose-response curves were generated for 60Co-γ-rays induced phospho-53 Binding Protein 1 (phospho-53BP1) foci at 1, 2, 4, 8, 16, and 24 h, post-irradiation for a dose range of 0.05–4 Gy using fluorescence microscopy. Following ISO recommendations, minimum detection limits (MDLs) were estimated to be 16, 18, 25, 40, 50, and 75 mGy for dose-response curves generated at 1, 2, 4, 8, 16, and 24 h post-irradiation. Colocalization and correlation of phospho-53BP1 and γH2AX were also measured in irradiated peripheral blood lymphocytes (PBLs) to gain dual confirmation. Comparative evaluation of the established curve was made by γH2AX-immunofluorescence, dicentric chromosome assay (DCA), and reciprocal translocation (RT) assays by reconstructing the dose of 6 dose-blinded samples. Coefficients of respective in-house established dose-response curves were employed to reconstruct the blind doses. Estimated doses were within the variation of 4.124%. For lower doses (0.052 Gy), phospho-53BP1 and γH2AX assays gave closer estimates with the variation of −4.1 to + 9% in comparison to cytogenetic assays, where variations were −8.5 to 24%. For higher doses (3 and 4 Gy), both the cytogenetic and immunofluorescence (phospho-53BP1 and γH2AX), assays gave comparable close estimates, with −11.3 to + 14.3% and −10.3 to −13.7%, variations, respectively. |
| first_indexed | 2024-04-13T11:14:01Z |
| format | Article |
| id | doaj.art-13eee02e88514b2a87239498199ba63a |
| institution | Directory Open Access Journal |
| issn | 2296-2565 |
| language | English |
| last_indexed | 2024-04-13T11:14:01Z |
| publishDate | 2022-08-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Public Health |
| spelling | doaj.art-13eee02e88514b2a87239498199ba63a2022-12-22T02:49:01ZengFrontiers Media S.A.Frontiers in Public Health2296-25652022-08-011010.3389/fpubh.2022.845200845200Establishment of in vitro Calibration Curve for 60Co-γ-rays Induced Phospho-53BP1 Foci, Rapid Biodosimetry and Initial Triage, and Comparative Evaluations With γH2AX and Cytogenetic AssaysRajesh Kumar Chaurasia0Rajesh Kumar Chaurasia1Kapil B. Shirsath2Utkarsha N. Desai3Nagesh N. Bhat4Nagesh N. Bhat5B. K. Sapra6B. K. Sapra7Radiological Physics and Advisory Division, Bhabha Atomic Research Centre (BARC), Mumbai, IndiaHomi Bhabha National Institute (HBNI), Mumbai, IndiaRadiological Physics and Advisory Division, Bhabha Atomic Research Centre (BARC), Mumbai, IndiaRadiological Physics and Advisory Division, Bhabha Atomic Research Centre (BARC), Mumbai, IndiaRadiological Physics and Advisory Division, Bhabha Atomic Research Centre (BARC), Mumbai, IndiaHomi Bhabha National Institute (HBNI), Mumbai, IndiaRadiological Physics and Advisory Division, Bhabha Atomic Research Centre (BARC), Mumbai, IndiaHomi Bhabha National Institute (HBNI), Mumbai, IndiaA rapid and reliable method for biodosimetry of populations exposed to ionizing radiation in the event of an incident or accident is crucial for initial triage and medical attention. DNA-double strand breaks (DSBs) are indicative of radiation exposure, and DSB-repair proteins (53BP1, γH2AX, ATM, etc.) are considered sensitive markers of DSB quantification. Phospho-53BP1 and γH2AX immunofluorescence technique serves as a sensitive, reliable, and reproducible tool for the detection and quantification of DSB-repair proteins, which can be used for biological dose estimations. In this study, dose-response curves were generated for 60Co-γ-rays induced phospho-53 Binding Protein 1 (phospho-53BP1) foci at 1, 2, 4, 8, 16, and 24 h, post-irradiation for a dose range of 0.05–4 Gy using fluorescence microscopy. Following ISO recommendations, minimum detection limits (MDLs) were estimated to be 16, 18, 25, 40, 50, and 75 mGy for dose-response curves generated at 1, 2, 4, 8, 16, and 24 h post-irradiation. Colocalization and correlation of phospho-53BP1 and γH2AX were also measured in irradiated peripheral blood lymphocytes (PBLs) to gain dual confirmation. Comparative evaluation of the established curve was made by γH2AX-immunofluorescence, dicentric chromosome assay (DCA), and reciprocal translocation (RT) assays by reconstructing the dose of 6 dose-blinded samples. Coefficients of respective in-house established dose-response curves were employed to reconstruct the blind doses. Estimated doses were within the variation of 4.124%. For lower doses (0.052 Gy), phospho-53BP1 and γH2AX assays gave closer estimates with the variation of −4.1 to + 9% in comparison to cytogenetic assays, where variations were −8.5 to 24%. For higher doses (3 and 4 Gy), both the cytogenetic and immunofluorescence (phospho-53BP1 and γH2AX), assays gave comparable close estimates, with −11.3 to + 14.3% and −10.3 to −13.7%, variations, respectively.https://www.frontiersin.org/articles/10.3389/fpubh.2022.845200/full53BP1 fociγH2AX focidicentrics and reciprocal translocationsrapid biodosimetryradiation induced DNA-DSB detectioncolocalization of γH2AX and 53BP1 foci |
| spellingShingle | Rajesh Kumar Chaurasia Rajesh Kumar Chaurasia Kapil B. Shirsath Utkarsha N. Desai Nagesh N. Bhat Nagesh N. Bhat B. K. Sapra B. K. Sapra Establishment of in vitro Calibration Curve for 60Co-γ-rays Induced Phospho-53BP1 Foci, Rapid Biodosimetry and Initial Triage, and Comparative Evaluations With γH2AX and Cytogenetic Assays Frontiers in Public Health 53BP1 foci γH2AX foci dicentrics and reciprocal translocations rapid biodosimetry radiation induced DNA-DSB detection colocalization of γH2AX and 53BP1 foci |
| title | Establishment of in vitro Calibration Curve for 60Co-γ-rays Induced Phospho-53BP1 Foci, Rapid Biodosimetry and Initial Triage, and Comparative Evaluations With γH2AX and Cytogenetic Assays |
| title_full | Establishment of in vitro Calibration Curve for 60Co-γ-rays Induced Phospho-53BP1 Foci, Rapid Biodosimetry and Initial Triage, and Comparative Evaluations With γH2AX and Cytogenetic Assays |
| title_fullStr | Establishment of in vitro Calibration Curve for 60Co-γ-rays Induced Phospho-53BP1 Foci, Rapid Biodosimetry and Initial Triage, and Comparative Evaluations With γH2AX and Cytogenetic Assays |
| title_full_unstemmed | Establishment of in vitro Calibration Curve for 60Co-γ-rays Induced Phospho-53BP1 Foci, Rapid Biodosimetry and Initial Triage, and Comparative Evaluations With γH2AX and Cytogenetic Assays |
| title_short | Establishment of in vitro Calibration Curve for 60Co-γ-rays Induced Phospho-53BP1 Foci, Rapid Biodosimetry and Initial Triage, and Comparative Evaluations With γH2AX and Cytogenetic Assays |
| title_sort | establishment of in vitro calibration curve for 60co γ rays induced phospho 53bp1 foci rapid biodosimetry and initial triage and comparative evaluations with γh2ax and cytogenetic assays |
| topic | 53BP1 foci γH2AX foci dicentrics and reciprocal translocations rapid biodosimetry radiation induced DNA-DSB detection colocalization of γH2AX and 53BP1 foci |
| url | https://www.frontiersin.org/articles/10.3389/fpubh.2022.845200/full |
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