M.Sc Thesis

M.Sc StudentHashem Mada
SubjectBiological Applications of Electron Spin Resonance
with High Spatial Resolution
DepartmentDepartment of Chemistry
Supervisor PROF. Aharon Blank
Full Thesis textFull thesis text - English Version


Oxygen (O2) plays a crucial role in many living processes. It is consumed by aerobic organisms to generate chemical energy, and it also serves as a regulatory molecule for important physiologic processes. For example, Reactive Oxygen Species (ROS) such as Superoxide is formed from the reduction of molecular oxygen, and its level at specific sites may affect cell pathogenesis and signaling. Any imbalance in tissue oxygen levels may affect metabolic homeostasis and lead to pathophysiological conditions. A complete understanding of oxygen-related process would require methods capable of quantifying the levels of tissue oxygenation as well as detecting ROS production with high accuracy and good spatial resolution.

This work is focused on the development and first usage of such O2 and ROS quantifying and mapping methods, based on Electron Spin Resonance (ESR). ESR is a widely used spectroscopic technique in the study of paramagnetic species, and has many applications in several fields of science. It can detect the concentrations of O2 and some types of ROS with high accuracy and specificity, but traditionally suffered from sensitivity and consequently, bad spatial resolution.

Recent progress in this technique has been aiming at alleviating these limitations and enabling the observation of small living samples with high sensitivity and high spatial resolution.

This study makes a further step in this methodological path by employing new resonators and imaging probes that are designed and developed especially to achieve this goal and to be able to make use ESR micro-imaging technique with small biological samples. We also present two applications for these newly developed methodologies, in the field of biology.

The first application uses ESR oximetry to accurately map the oxygen levels in spheroids of cancer cells in a three dimensional manner.

The second application employs the spin probe technique to detect the production of superoxide in the roots of the delicate plant Arabidopsis thaliana, following some stress signal. Extracellular accumulation of superoxide was successfully detected at specific sites of the root.