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Calibration of the γ-H2AX DNA double strand break focus assay for internal radiation exposure of blood lymphocytes.

Eberlein U, Peper M, Fernández M, Lassmann M, Scherthan H - PLoS ONE (2015)

Bottom Line: The measured blood doses in our samples ranged from 6 to 95 mGy.A linear relationship was found between the number of DSB-marking foci/nucleus and the absorbed dose to the blood for both radionuclides studied.There were only minor nuclide-specific intra- and inter-subject deviations.

View Article: PubMed Central - PubMed

Affiliation: Department of Nuclear Medicine, University of Würzburg, Würzburg, Germany.

ABSTRACT
DNA double strand break (DSB) formation induced by ionizing radiation exposure is indicated by the DSB biomarkers γ-H2AX and 53BP1. Knowledge about DSB foci formation in-vitro after internal irradiation of whole blood samples with radionuclides in solution will help us to gain detailed insights about dose-response relationships in patients after molecular radiotherapy (MRT). Therefore, we studied the induction of radiation-induced co-localizing γ-H2AX and 53BP1 foci as surrogate markers for DSBs in-vitro, and correlated the obtained foci per cell values with the in-vitro absorbed doses to the blood for the two most frequently used radionuclides in MRT (I-131 and Lu-177). This approach led to an in-vitro calibration curve. Overall, 55 blood samples of three healthy volunteers were analyzed. For each experiment several vials containing a mixture of whole blood and radioactive solutions with different concentrations of isotonic NaCl-diluted radionuclides with known activities were prepared. Leukocytes were recovered by density centrifugation after incubation and constant blending for 1 h at 37°C. After ethanol fixation they were subjected to two-color immunofluorescence staining and the average frequencies of the co-localizing γ-H2AX and 53BP1 foci/nucleus were determined using a fluorescence microscope equipped with a red/green double band pass filter. The exact activity was determined in parallel in each blood sample by calibrated germanium detector measurements. The absorbed dose rates to the blood per nuclear disintegrations occurring in 1 ml of blood were calculated for both isotopes by a Monte Carlo simulation. The measured blood doses in our samples ranged from 6 to 95 mGy. A linear relationship was found between the number of DSB-marking foci/nucleus and the absorbed dose to the blood for both radionuclides studied. There were only minor nuclide-specific intra- and inter-subject deviations.

No MeSH data available.


Related in: MedlinePlus

Average RIF/cell as a function of the absorbed dose.Graph showing the results of the individual measurements for our test persons’ blood samples (TP1-TP3) treated with I-131 (I) and Lu-177 (Lu). The error bars represent standard deviation of each single lymphocyte sample, assuming a Poisson distribution.
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pone.0123174.g002: Average RIF/cell as a function of the absorbed dose.Graph showing the results of the individual measurements for our test persons’ blood samples (TP1-TP3) treated with I-131 (I) and Lu-177 (Lu). The error bars represent standard deviation of each single lymphocyte sample, assuming a Poisson distribution.

Mentions: We determined the RIF per cell as a function of the absorbed dose to the blood for each test person and measurement only minor nuclide-specific, intra- and inter-subject deviations between I-131 and Lu-177 were observed (Fig 2).


Calibration of the γ-H2AX DNA double strand break focus assay for internal radiation exposure of blood lymphocytes.

Eberlein U, Peper M, Fernández M, Lassmann M, Scherthan H - PLoS ONE (2015)

Average RIF/cell as a function of the absorbed dose.Graph showing the results of the individual measurements for our test persons’ blood samples (TP1-TP3) treated with I-131 (I) and Lu-177 (Lu). The error bars represent standard deviation of each single lymphocyte sample, assuming a Poisson distribution.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4390303&req=5

pone.0123174.g002: Average RIF/cell as a function of the absorbed dose.Graph showing the results of the individual measurements for our test persons’ blood samples (TP1-TP3) treated with I-131 (I) and Lu-177 (Lu). The error bars represent standard deviation of each single lymphocyte sample, assuming a Poisson distribution.
Mentions: We determined the RIF per cell as a function of the absorbed dose to the blood for each test person and measurement only minor nuclide-specific, intra- and inter-subject deviations between I-131 and Lu-177 were observed (Fig 2).

Bottom Line: The measured blood doses in our samples ranged from 6 to 95 mGy.A linear relationship was found between the number of DSB-marking foci/nucleus and the absorbed dose to the blood for both radionuclides studied.There were only minor nuclide-specific intra- and inter-subject deviations.

View Article: PubMed Central - PubMed

Affiliation: Department of Nuclear Medicine, University of Würzburg, Würzburg, Germany.

ABSTRACT
DNA double strand break (DSB) formation induced by ionizing radiation exposure is indicated by the DSB biomarkers γ-H2AX and 53BP1. Knowledge about DSB foci formation in-vitro after internal irradiation of whole blood samples with radionuclides in solution will help us to gain detailed insights about dose-response relationships in patients after molecular radiotherapy (MRT). Therefore, we studied the induction of radiation-induced co-localizing γ-H2AX and 53BP1 foci as surrogate markers for DSBs in-vitro, and correlated the obtained foci per cell values with the in-vitro absorbed doses to the blood for the two most frequently used radionuclides in MRT (I-131 and Lu-177). This approach led to an in-vitro calibration curve. Overall, 55 blood samples of three healthy volunteers were analyzed. For each experiment several vials containing a mixture of whole blood and radioactive solutions with different concentrations of isotonic NaCl-diluted radionuclides with known activities were prepared. Leukocytes were recovered by density centrifugation after incubation and constant blending for 1 h at 37°C. After ethanol fixation they were subjected to two-color immunofluorescence staining and the average frequencies of the co-localizing γ-H2AX and 53BP1 foci/nucleus were determined using a fluorescence microscope equipped with a red/green double band pass filter. The exact activity was determined in parallel in each blood sample by calibrated germanium detector measurements. The absorbed dose rates to the blood per nuclear disintegrations occurring in 1 ml of blood were calculated for both isotopes by a Monte Carlo simulation. The measured blood doses in our samples ranged from 6 to 95 mGy. A linear relationship was found between the number of DSB-marking foci/nucleus and the absorbed dose to the blood for both radionuclides studied. There were only minor nuclide-specific intra- and inter-subject deviations.

No MeSH data available.


Related in: MedlinePlus