Sensitivity of Human Lymphocytes to Sensitivity of Human Lymphocytes to Low-Dose RadiationLow-Dose Radiation

Gordon K. Livingston, PhDGordon K. Livingston, PhD

Technical Director, Cytogenetic Biodosimetry LaboratoryTechnical Director, Cytogenetic Biodosimetry Laboratory

Radiation Emergency Assistance Center Training SiteRadiation Emergency Assistance Center Training Site

Oak Ridge Insitute for Science and EducationOak Ridge Insitute for Science and Education

Oak Ridge, TN 37831Oak Ridge, TN 37831

Gordon.Livingston@orise.orau.gov

http://orise.orau.gov/reactshttp://orise.orau.gov/reacts

Cytogenetic BiodosimetryCytogenetic Biodosimetry

Cytogenetic analysis provides a method to estimate radiation dose to individuals exposed to ionizing radiation when

physical dosimetry devices are not available.

LymphocytesLymphocytes

Lymphocytes serve as ideal biological dosimeters because they: 1) are very sensitive to radiation,

2) are readily available in routine blood samples,

3) circulate throughout the body,

4) can be easily cultured and induced to divide in vitro

5) respond consistently to ionizing radiation exposure by expressing chromosomal DNA damage in a dose-dependent manner.

Image of Normal Metaphase Spread

Dicentric ChromosomeDicentric Chromosome

Dicentric Chromosome AssayDicentric Chromosome Assay

Low background rate (1-2 per 1,000 cells) Independent of age and gender Sensitivity range (0.2 – 5 Gy) Specificity to radiation is high Consistent radiation dose-response Proven in accidents over four decades Considered the gold standard

Multiple Dicentric ChromosomesMultiple Dicentric Chromosomes

Typical Dose Response CurvesTypical Dose Response Curves

Y=D+D2

Quadratic (linear-quadratic)

Y=DLinear

Dic

entr

ics/

cell

Dose [Gy]

The Lymphocyte Micronucleus The Lymphocyte Micronucleus Test as a Radiation BiodosimeterTest as a Radiation Biodosimeter

LyLLy

Lymphocyte in Normal Anaphase

Lymphocyte in Abnormal Anaphase

Micronucleus

X-Ray Therapy MachineX-Ray Therapy Machine

X-ray Exposure ConditionsX-ray Exposure Conditions

Westinghouse Orthovoltage Therapy Unit

78 kV, 5 mA, 2mm Al added filtration

output measured at 19.8 cGy/min at 50 cm

30 cm field size with +/- 10 % uniformity

graded doses of 0, 5, 10, 15, 20 & 25 cGy

10 ml media (3-4 x 105 cells/ml) in 25 cm2 culture flasks at 37 C for 72 hr

Micronucleus Dose ResponseMicronucleus Dose Response

The Micronucleus Test as a The Micronucleus Test as a Biological DosimeterBiological Dosimeter

Linear Dose Response to X-rays

y = 2.783 x + 3.714

35.5 = 2.783 x + 3.714

X = 11.4 c Gy

NCRP Dose to red marrow is 0.2 m rad / micro Ci

48,000 micro Ci x 0.2 m rad = 9,600 m rad

= 9.6 rad (9.6 c Gy)

Green 3, 5 & 6

Red 1, 2 & 4

FISH - Total Translocations by Group

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FISH-Exposed; High-Dose Group (n=15)

FISH-Unexposed Controls (n=15)

I-131 Internal Exposure

Partial Body X-ray Exposure

Scoring Dicentrics with New Technologies

80 Slide Feeder

Digital Camera and Images

Automated slide scanning stage.

Automated Cytogenetic Workstation

Digital images are used for scoring chromosomes aberrations

(dicentrics).

Scoring Dicentrics with New Technologies

80 Slide Feeder

Digital Camera and Images

Automated slide scanning stage.

Automated Cytogenetic Workstation

Digital images are used for scoring chromosomes aberrations

(dicentrics).

Cytogenetic BiodosimetryCytogenetic Biodosimetry

Limitations– Large event could overwhelm resources– Labor intensive– Samples require shipment

Advantages– Proven method for dose estimation – Amenable to web-based image scoring– Existing resource of clinical laboratories

Cytogenetic Dosimetry Cytogenetic Dosimetry Capacity WorldwideCapacity Worldwide

Cytogenetic labs and regional networks already

collaborating with WHO:– North-American– Latin American (with IAEA support)– European– Scandinavian– Eastern Europe: RF and Ukraine– Western-Pacific: Japan, S. Korea, Australia

Need to integrate and coordinate these networks globally

Need a clear concept of operations for emergencies and in

"quiet" time to make the most of these labs joint capabilities

ConsortiumConsortium

REAC/TS CBL

Secure Central Server (Access Limited)

Lab in Consortium (Metafer Software)

Satellite Scoring Station (Metafer Software)

Lab in Consortium (Metafer System)

The re-establishment of the REAC/TS-Cytogenetic The re-establishment of the REAC/TS-Cytogenetic Biodosimetry Laboratory, operated by the Oak Biodosimetry Laboratory, operated by the Oak Ridge Institute for Science and Education Ridge Institute for Science and Education (ORISE), was supported by: (ORISE), was supported by:

DOE-NNSA, DOE-NNSA, DOE-EH, and DOE-EH, and Nuclear Regulatory Commission.Nuclear Regulatory Commission.

Establishing a Web-Based International Radiation Establishing a Web-Based International Radiation Cytogenetic Biodosimetry Consortium at REAC/TS is Cytogenetic Biodosimetry Consortium at REAC/TS is

being funded by DOE-NNSA.being funded by DOE-NNSA.

QuestionsQuestions