Ms. Angelina I. Dolgikh Profile

Angelina I. Dolgikh

at Orel State Univ

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Proceedings Article | 11 August 2023 Paper

Determination of the physiological state of cells by differences in FAD fluorescence intensity

A. Dolgikh, E. Bryanskaya, A. Yu. Vinokurov, A. Dunaev

Proceedings Volume 12627, 126272L (2023) https://doi.org/10.1117/12.2670961

KEYWORDS: Fluorescence intensity, Autofluorescence, Signal intensity, Fluorescence, Skin, Safety, Pathology, Oxidation, Diseases and disorders

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This work considers the measurement of FAD fluorescence intensity as a method for the safety, simple and real-time detection of pathological cells and informative value of this approach. FAD participates in essential processes such as fatty acid oxidation, the Krebs cycle and other redox reactions. According to literature, cells in different physiological states have different levels of FAD intensity in green-blue spectrum. Hence, it is highly relevant to determine the physiological state of cells by the difference in FAD signal intensity. The study was realized with skin fibroblasts as a model object. On the first stage of experiments 20-days cells cultured on a pre-marked coverslips were divided into two subgroups on the basis of the autofluorescence signal intensity. The first subgroup included cells with a high autofluorescence signal (presumably senescent or pathological), and the second – cells and low one. During subsequent experiments after 24 hours necrotic cells were analyzed in a culture using Hoechst 33342 and propidium iodide in two subgroups separately. According to the results, over 50% of cells with high autofluorescence intensity were identified as necrotic, that can subsequently be used for early diagnosis of various pathologies states. Thus, this study, with its advantages such as non-invasiveness, high sensitivity and biosafety, shows the possibility of early diagnosis of various diseases by measuring the fluorescence signal of FAD and finding cells with high fluorescence levels, which are mostly necrotic.

Proceedings Article | 4 May 2021 Paper

Measurements of mitochondrial NADH pool and NADH production rate in acute brain slices and primary cell cultures using live cell imaging

Angelina Dolgikh, Olga Stelmashchuk, Andrey Vinokurov, Evgeny Zherebtsov, Andrey Abramov

Proceedings Volume 11845, 118450L (2021) https://doi.org/10.1117/12.2590804

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The paper presents results of the estimation of bioenergetic parameters in acute slices of brain rat by measuring mitochondrial pool and rate of NADH production using fluorescence imaging. NAD+ and NADH play crucial roles in mitochondrial energy metabolism due to their participation in the tricarboxylic acid cycle and in electron transport chain. That is why it is essential to know variability of its content in cells. In the study we used an approach based on the registration of auto-fluorescence of NADH. An experimental setup was designed and assembled for the imaging of the acute brain slices but also capable of measuring brain cell cultures. BDL-SMN-375 laser (Becker&Hickl, Germany) was used to excite fluorescence at the wavelength of 375 nm. The parameters of NADH content in both acute brain slices and cell cultures of midbrain and cortex were estimated. The obtained results can be of interest for better understanding of development of neurodegenerative diseases of mitochondrial disfunction origin.

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