Academic Journal
Development of a high-throughput γ-H2AX assay based on imaging flow cytometry
| العنوان: | Development of a high-throughput γ-H2AX assay based on imaging flow cytometry |
|---|---|
| المؤلفون: | Younghyun Lee, Qi Wang, Igor Shuryak, David J. Brenner, Helen C. Turner |
| المصدر: | Radiation Oncology, Vol 14, Iss 1, Pp 1-10 (2019) |
| بيانات النشر: | BMC, 2019. |
| سنة النشر: | 2019 |
| المجموعة: | LCC:Medical physics. Medical radiology. Nuclear medicine LCC:Neoplasms. Tumors. Oncology. Including cancer and carcinogens |
| مصطلحات موضوعية: | Imaging flow cytometry, DNA repair kinetics, Human lymphocytes, High throughput, Radiation sensitivity, Ionizing radiation, Medical physics. Medical radiology. Nuclear medicine, R895-920, Neoplasms. Tumors. Oncology. Including cancer and carcinogens, RC254-282 |
| الوصف: | Abstract Background Measurement of γ-H2AX foci levels in cells provides a sensitive and reliable method for quantitation of the radiation-induced DNA damage response. The objective of the present study was to develop a rapid, high-throughput γ-H2AX assay based on imaging flow cytometry (IFC) using the ImageStream®X Mk II (ISX) platform to evaluate DNA double strand break (DSB) repair kinetics in human peripheral blood cells after exposure to ionizing irradiation. Methods The γ-H2AX protocol was developed and optimized for small volumes (100 μL) of human blood in Matrix™ 96-tube format. Blood cell lymphocytes were identified and captured by ISX INSPIRE™ software and analyzed by Data Exploration and Analysis Software. Results Dose- and time-dependent γ-H2AX levels corresponding to radiation exposure were measured at various time points over 24 h using the IFC system. γ-H2AX fluorescence intensity at 1 h after exposure, increased linearly with increasing radiation dose (R 2 = 0.98) for the four human donors tested, whereas the dose response for the mean number of γ-H2AX foci/cell was not as robust (R 2 = 0.81). Radiation-induced γ-H2AX levels rapidly increased within 30 min and reached a maximum by ~ 1 h, after which time there was fast decline by 6 h, followed by a much slower rate of disappearance up to 24 h. A mathematical approach for quantifying DNA repair kinetics using the rate of γ-H2AX decay (decay constant, Kdec), and yield of residual unrepaired breaks (Fres) demonstrated differences in individual repair capacity between the healthy donors. Conclusions The results indicate that the IFC-based γ-H2AX protocol may provide a practical and high-throughput platform for measurements of individual global DNA DSB repair capacity which can facilitate precision medicine by predicting individual radiosensitivity and risk of developing adverse effects related to radiotherapy treatment. |
| نوع الوثيقة: | article |
| وصف الملف: | electronic resource |
| اللغة: | English |
| تدمد: | 1748-717X |
| Relation: | http://link.springer.com/article/10.1186/s13014-019-1344-7; https://doaj.org/toc/1748-717X |
| DOI: | 10.1186/s13014-019-1344-7 |
| URL الوصول: | https://doaj.org/article/2e3181759daa4be1a3fa693ad277afa2 |
| رقم الانضمام: | edsdoj.2e3181759daa4be1a3fa693ad277afa2 |
| قاعدة البيانات: | Directory of Open Access Journals |
Full Text Finder | تدمد: | 1748717X |
|---|---|
| DOI: | 10.1186/s13014-019-1344-7 |