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Probing living cells by terahertz Attenuated Total Reflection : permeabilization dynamics of the cell membrane

Abstract : Terahertz radiation is located in the electromagnetic range between far infrared and microwaves, corresponding to frequencies between 0.1 and 10 THz. This spectral range is currently largely under-exploited, but its application to the study of biological objects has already shown a strong potential, in the detection of skin cancer, ion flow monitoring or biosensors. In the field of biology, which is of particular interest to us here, the terahertz range makes it possible to quantify and discriminate solutes of biological interest thanks to the interaction with low-frequency modes of liquid water, and thus to study biomolecules, microorganisms and cells in their physiological environment. The first part of this thesis work consisted in studying the dynamics of membrane permeabilization of living cells by attenuated total reflection (ATR) with our device based on a femtosecond laser and the generation of ultrashort terahertz pulses. Monolayers of MDCK epithelial cells were exposed to varying concentrations of saponin, a detergent that digs holes in the cell membrane. The dynamics obtained were then compared to a theoretical model describing the physical behavior of the cell layer, taking into account the diffusion of detergent molecules and the physical characteristics of the membrane. The good agreement between experiment and theory indicates that membrane permeabilization is limited mainly by the diffusion of detergent molecules and their binding to the membrane.In a second part, we developed a completely new system based on a continuous terahertz QCL source at 2.5 THz. This new instrument is based on a very simplified design, with a single ATR prism and a single detector, and on the dual modulation of the terahertz beam using a mechanical chopper. This chopper synchronizes the dual modulation and defines the measurement and reference zones. The long-term stability of this device has been greatly enhanced by the precise control of temperature and humidity inside the device. Performance is excellent in both the short and long term. A signal-to-noise ratio of 30 dB is achieved over 300ms, and remains above 30 dB for several hours. In addition, a theoretical and experimental study has been carried out to calibrate the instrument. Thus, the ATR reflection coefficients of several solutions of biological interest (ions, sugars and proteins) were obtained over a wide range of concentrations. A sensitivity at least 20 times higher than that of the existing literature was thus obtained. Thanks to this new high-performance system, we studied the dynamics of membrane permeabilization following the action of photodynamic therapy (PDT). The first results showed that the encapsulation of photosensitizers by micellar vectors significantly improves the efficiency of PDT.
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Submitted on : Tuesday, February 23, 2021 - 4:59:40 PM
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Xiujun Zheng. Probing living cells by terahertz Attenuated Total Reflection : permeabilization dynamics of the cell membrane. Biological Physics [physics.bio-ph]. Institut Polytechnique de Paris, 2020. English. ⟨NNT : 2020IPPAX079⟩. ⟨tel-03150356⟩

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