Use Of Atomic Force Microscopy Research Articles

Abstract PS03-08: Spatial biomechanics determines fate in breast cancer

Abstract Background: With the recent advancements of spatial omics, it has become increasingly recognized that spatial analysis the of tumor architecture is key to decipher the heterogenous intratumoral relationships between different tumor components and provide better understanding of cancer therapy resistance, as well as help to identify potential targets for personalized therapy. The challenge of the discovery and implementation of such biomarkers still lays in the technological and methodological difficulties, and their translation into clinical practice. In parallel a major advancement in cancer research has been the recognition of the importance of cancer biomechanical properties in cancer progression, metastasis, and therapy response. Methods: We developed a new computational platform to identify biomechanical drivers of cancer outcome from spatial omics data. We used it to identify recurring spatial patterns of these markers within the tumor microenvironment, and define the spatial scale of heterogeneity of such patterns. Our dataset consists in 700 primary breast cancer baseline samples analyzed with two 30-marker imaging mass cytometry panels to identify cancer, immune, and stromal cells and structures and their biomechanical states. Our patient cohort includes 20+ years of follow up, with up to 6 samples per patient from 64 patients alive 12 years post diagnosis, and 49 patients lost to breast cancer deaths. Patients were treated with surgery, radiation, chemotherapy, endocrine therapy, or combinations of these post-surgery. Based on this approach, we derived a new three-parameter quantitative metric of preferential spatial co-localization between different cells or structures, and we use this metric to stratify patients into responders and non-responders with the aid of single and multivariate survival analysis. Finally, we include a correlation with Atomic Force Microscopy to identify the mechanical signature of these driving patterns in breast cancer. Results: Our work enabled the identification of tumor-immune-stromal spatial patterns that drive breast cancer outcome and their spatial scale. Among our results, we were able to further define spatial regions of hypoxia-driven EMT. We found these regions to be usually on the scale of 50 mm radius, and enriched in presence of unspecified immune cells. They are also more frequent in ER+ areas, with high NaKATPase activity. Our analysis also shed light on the controversy on the role of fibroblasts. We have found that the presence of a strong, spatially structured stroma, in fibroblast-rich regions is a strong predictor of positive outcome. Similarly, in our analysis collagen cross-linking emerges as a positive control factor in Vimentin-rich microenvironments, offering new interpretation to the role of the ECM structure. Importantly, our first principle approach allows for the identification of driving structures beyond heterogeneity, and thus enables easy extension to additional datasets. Finally, when correlating these patterns with Atomic force microscopy measurements, we can clearly see that patterns predictive of poor survival present a clear heterogeneous stiffness signature, as opposed to a very homogenous good survival pattern signature. Conclusion: These results confirm the importance of spatial distribution of biomechanical drivers in cancer, offering new avenues of physics-based therapeutic targets. Our results also offer a solid base to inform machine learning algorithms on how to significantly parametrize spatial patterns in breast cancer for clinical significance. Furthermore, we demonstrate for the first time the use of Atomic Force Microscopy as a single biomarker for these patterns in breast cancer with a direct correlation based on pathology matching. Citation Format: Sara Nizzero, Maria Pelaez Soni, Gregory Zaugg, Mariam Gachechiladze, Yitian Xu, Licheng Zhang, Junjun Zheng, Brian Menegaz, Lee B Jordan, Colin A Purdie, Philip R Quinlan, Chandandeep Nagi, Karla A Sepulveda, Philipp Oertle, Tobias A Appenzeller, Marko Loparic, Zhihui Wang, Shu-Hsia Chen, Vittorio Cristini, Marija Plodinec, Alastair Thompson. Spatial biomechanics determines fate in breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PS03-08.

A DNA Force Circuit for Exploring Protein‐Protein Interactions at the Single‐Molecule Level†

Comprehensive SummaryProtein‐protein interactions (PPIs) play a crucial role in drug discovery and disease treatment. However, the development of effective drugs targeting PPIs remains challenging due to limited methodologies for probing their spatiotemporal anisotropy. Here, we propose a single‐molecule approach using a unique force circuit to investigate PPI dynamics and anisotropy under mechanical forces. Unlike conventional techniques, this approach enables the manipulation and real‐time monitoring of individual proteins at specific amino acids with defined geometry, offering insights into molecular mechanisms at the single‐molecule level. The DNA force circuit was constructed using click chemistry conjugation methods and genetic code expansion techniques, facilitating orthogonal conjugation between proteins and nucleic acids. The SET domain of the MLL1 protein and the tail of histone H3 were used as a model system to demonstrate the application of the DNA force circuit. With the use of atomic force microscopy and magnetic tweezers, optimized assembly procedures were developed. The DNA force circuit provides an exceptional platform for studying the anisotropy of PPIs and holds promise for advancing drug discovery research targeted at PPIs.

Characterization of the mechanical properties of the cortex region of human hair fibers by multiparametric atomic force microscopy mapping

We show the benefit of the use of atomic force microscopy (AFM) in spectroscopy force mode (FV: force volume) for evaluation of the cosmetic active effectiveness in improving the mechanical properties of human hair fibers cortex region. For this, we characterized human hair fibers without and with chemical damage caused by bleaching process. Fiber and resin (embedding material) data were obtained simultaneously in the mapping in order to have the resin data as a reference to ensure a coherent comparison between data from the different fiber groups. Our AFM results, which were evaluated using statistical tests, demonstrated the degradation of fibers after bleaching, corroborating the findings of transmission electron microscopy analysis and the effectiveness of a cosmetic active ingredient in improving the Young's modulus (elastic modulus) (E) of the damaged fibers. We also found a radial decrease in the natural logarithm of Young's modulus ln(E) along the cross-section of the active group fiber, which is compatible with confocal Raman spectroscopy analysis by other authors, demonstrating variation of the active permeation with depth. We note that Young's modulus was also determined by a tensile tester (macro-scale technique), in which it was not possible to obtain statistically significant differences between the groups, evidencing the advantage of the FV-AFM analysis. We also found an increase in ln(E) accompanied by a decrease in maximum adhesion force between tip and sample (negative Pearson correlation coefficient). This result can be explained by the fact that structures composed of hydrophobic components have a higher Young's modulus than structures composed of hydrophilic components.•Bleaching damage and cosmetic hair treatment assessed by AFM, TEM, and tensile tester.•Young's modulus by AFM nanoindentation of hair fibers monitored by sample standard.•Young's modulus changes radially along the cross-section due to the cosmetic active.•AFM data show statistically significant differences among sample groups.•Tensile tester was not able to show statistically significant differences.

Advances in the Use of Atomic Force Microscopy as a Diagnostic Tool for Solar Cells Characterization: From Material Design to Device Applications

Considerable efforts in search for an effective characterization technique for photovoltaic devices with utmost precision is on the increase. For precise analysis and tailoring of device performance, a reliable technique is vital. Atomic force microscopy is one of the leading surface analysis techniques of choice for probing surface patterns in a variety of materials with atomic precision using a cantilever. It has evolved as a reliable technique for the investigation of subatomic scale properties of materials such as photocurrent heterogeneity, electromechanical response, charge distribution, molecular weight effects, and many other material parameters. The integration of artificial intelligence hybrid algorithms in atomic force microscope for optoelectronic device fabrication and characterization has increasingly emerged to be desirable due to its reliability and effectiveness in achieving high image resolution, automated analysis, actuation, and the coupling of manufactured units with a precision down to atomic units. In this review, an investigation of topical developments in the use of atomic force microscopy as a diagnostic tool for solar cells characterization is presented with special focus on polymer solar cells, perovskite solar cells, quantum dots‐sensitized solar cells, dye‐sensitized solar cells, fullerene‐based solar cells, III‐V‐based solar cells, and silicon‐based solar cells.

Use of atomic force microscopy for characterization of model membranes and cells

Background: To provide a fundamental understanding of the potential and use of atomic force microscopy (AFM) in medicine and the life sciences, this work presents a thorough description of imaging and non-imaging atomic force microscopy modes for characterizing model membranes and cells at the nanoscale. Methods: The imaging and non-imaging AFM modes are described with examples in terms of the characterization of topographic, morphological, and nanomechanical sample properties. Results: AFM imaging of supported lipid bilayers (SLBs) revealed the effects of temperature and medium composition on SLB topography in the gel and fluid phases, and on the bilayer thickness. Non-imaging AFM showed the strengthening of the SLB in both phases by the ion binding process.Imaging of neuronal and neuroblastoma cells with and without treatment revealed morphological changes in shape, volume, roughness, and Feret dimension. Non-imaging AFM showed the change in cell elasticity induced by the treatment with H2O2 with and without quercetin and by the treatment with copper and myricetin. The measurements of cells elasticity revealed a reorganization of the cytoskeleton and filament structures. Conclusions: Diverse applications of imaging and non-imaging AFM can provide important information about the underlying processes in biologically relevant systems. AFM, as a complementary technique to other biomedical methods, allows screening and monitoring of physiological changes at the nanoscale.

Polyurethane-Based Nanocomposites for Regenerative Therapies of Cancer Skin Surgery with Low Inflammatory Potential to Healthy Fibroblasts and Keratinocytes In Vitro.

Nanocomposites based on thermoplastic polyurethanes (TPUs) filled with halloysite nanotubes (HNTs) were studied for their physicochemical and biological properties. Nanocomposites containing halloysite nanotube filler contents of 1 and 2% (E+1 and E+2), respectively, were obtained by extrusion. The newly formed E+1 and E+2 nanomaterials exhibited better flexibility and similar thermal properties compared to neat polyurethane. The use of atomic force microscopy (AFM) and differential scanning calorimetry (DSC) thermogram analysis showed that the distribution of halloysite nanotubes in the polymer matrix is more evenly dispersed in the E+1 nanomaterial, where the grains in the E+2 nanomaterial have a greater tendency to form agglomerates. Mechanical tests have shown that nanocomposites with the addition of HNT are characterized by a higher stress at break and elongation at break compared to neat TPU. The results of cytotoxicity tests suggest that the nanocomposite materials express lower toxicity to normal HaCaT and NHDF than to cancer Me45 cells. Further studies showed that the tested materials induced the expression of proinflammatory interleukins IL6 and IL8 in normal cells, but their overexpression in the cancer cell line resulted in cytostatic effects and proliferation reduction. Such a conclusion suggests the possible application of tested materials for regenerative therapies in cancer surgeries.

Atomic Force Microscopy of Phytosterol Based Edible Oleogels.

This work reviews the use of atomic force microscopy (AFM) as a tool to investigate oleogels of edible triglyceride oils. Specific attention is given to those oleogels based on phytosterols and their esters, a class of material the authors have studied extensively. This work consists of a summary of the role of AFM in imaging edible oleogels, including the processing and preparation steps required to obtain high-quality AFM images of them. Finally, there is a comparison between AFM and other techniques that may be used to obtain structural information from oleogel samples. The aim of this review is to provide a useful introduction and summary of the technique for researchers in the fields of gels and food sciences looking to perform AFM measurements on edible oleogels.

Use of Atomic Force Microscopy in UVB-Induced Chromosome Damage Provides Important Bioinformation for Cell Damage Assessment.

The chromosomal structure derived from UVB-stimulated HaCaT cells was detected by atomic force microscopy (AFM) to evaluate the effect of UVB irradiation. The results showed that the higher the UVB irradiation dose, the more the cells that had chromosome aberration. At the same time, different representative types of chromosome structural aberrations were investigated. We also revealed damage to both DNA and cells under the corresponding irradiation doses. It was found that the degree of DNA damage was directly proportional to the irradiation dose. The mechanical properties of cells were also changed after UVB irradiation, suggesting that cells experienced a series of chain reactions from inside to outside after irradiation. The high-resolution imaging of chromosome structures by AFM after UVB irradiation enables us to relate the damage between chromosomes, DNA, and cells caused by UVB irradiation and provides specific information on genetic effects.

Morphological Investigation of Protein Crystals by Atomic Force Microscopy

In this review, we discuss the progress in the investigation of macromolecular crystals obtained through the use of atomic force microscopy (AFM), a powerful tool for imaging surfaces and specimens at high resolution. AFM enables the visualization of soft samples at the nanoscale and can provide precise visual details over a wide size range, from the molecular level up to hundreds of micrometers. The nonperturbative nature, the ability to scan in a liquid environment, and the lack of need for freezing, fixing, or staining make AFM a well-suited tool for studying fragile samples such as macromolecular crystals. Starting from the first morphological investigations revealing the surface morphology of protein crystals, this review discusses the achievements of AFM in understanding the crystal growth processes, both at the micro- and nanoscale. The capability of AFM to investigate the sample structure at the single molecular level is analyzed considering in-depth the structure of S-layers. Lastly, high-speed atomic force microscopy (HS-AFM) is discussed as the evolution to overcome the limitations of low imaging speed, allowing for the observation of molecular dynamics and weakly adsorbed, diffusing molecules. HS-AFM has provided intuitive views and directly visualized phenomena that were previously described indirectly, answering questions that were challenging to address using other characterization methods.

Myelination dictates axonal viscoelasticity.

During development, dramatic changes in myelination, growth of neural networks and changes in grey-to-white matter ratio build up the astonishingly plastic brain of a child. The progressive increase in myelination insulates the nervous system, which, in turn, modifies the mechanical microenvironment of the brain spatiotemporally. A growing body of evidence demonstrates the role of mechanical forces in growth, differentiation, maturation and electrical properties of neurons. However, due to limitations in imaging resolution, the exact relationship between myelination, axonal organization and the mechanical properties of nerves at the cellular level is still unknown. Here, we propose a novel approach to study the direct relationship between axonal viscoelasticity with changing fibre anisotropy and myelination during development. With the use of atomic force microscopy (AFM) with in situ fluorescent imaging of the primary neuron-oligodendrocyte co-cultures, we found that as axons are progressively myelinated in vitro, their stiffness increases. Direct quantification of myelin along axons using immunofluorescence also demonstrated a positive correlation between increased myelination over time and increased axonal stiffness (p = .001). Notably, AFM measurements along a single axon showed that the Young's modulus measured across myelinated regions were significantly higher than those of adjacent unmyelinated segments at all time points (p < .0001). Force-relaxation analysis also demonstrated that myelin sheath dominates the regulation of viscoelasticity of axons temporally. Collectively, our findings indicate a direct link between myelination, axonal orientation and viscoelasticity, providing important insights about the mechanical environment in the paediatric brain, with direct implications for our understanding of developmental brain disorders and paediatric brain injury.

Mechanical and magnetic properties of Mg-Ni hydrosilicate nanoscrolls as study objects for atomic force microscopy

In hydrosilicates with the structure of chrysotile, pecoraite, and halloysite, elastic strains in the composite hydrosilicate layer are compensated by folding into nanoscrolls. A wide range of such particles with different compositions can be produced by hydrothermal synthesis, which, together with the structural and morphological characteristics, makes them promising adsorbents, capsules, catalysts, and reinforcing components of composite materials. For such potential applications as magnetically controlled adsorbents and catalysts, ferromagnetic chemical elements are included in the composition of the nanoscrolls. Studies of the individual nanoscrolls require special approaches. This article discusses the use of atomic force microscopy for such studies.

Nanomechanical Signatures of Extracellular Vesicles from Hematologic Cancer Patients Unraveled by Atomic Force Microscopy for Liquid Biopsy.

Cells release extracellular vesicles (EVs) as the carriers for intercellular communications to regulate life activities. Particularly, it is increasingly apparent that mechanical forces play an essential role in biological systems. The nanomechanical properties of EVs and their dynamics in cancer development are still not fully understood. Herein, with the use of atomic force microscopy (AFM), the nanomechanical signatures of EVs from the liquid biopsies of hematologic cancer patients were unraveled. Single native EVs were probed by AFM under aqueous conditions. The elastic and viscous properties of EVs were measured and visualized to correlate EV mechanics with EV geometry. Experimental results remarkably reveal the significant differences in EV mechanics among multiple myeloma patients, lymphoma patients, and healthy volunteers. The study unveils the unique nanomechanical signatures of EVs in hematologic cancers, which will benefit the studies of liquid biopsies for cancer diagnosis and prognosis with translational significance.

Use of atomic force microscopy to view ultrastructure of the fiber cell wall in Phyllostachys edulis culms

Use of atomic force microscopy to view ultrastructure of the fiber cell wall in Phyllostachys edulis culms

Machine learning of atomic force microscopy images of organic solar cells.

The bulk heterojunction structures of organic photovoltaics (OPVs) have been overlooked in their machine learning (ML) approach despite their presumably significant impact on power conversion efficiency (PCE). In this study, we examined the use of atomic force microscopy (AFM) images to construct an ML model for predicting the PCE of polymer : non-fullerene molecular acceptor OPVs. We manually collected experimentally observed AFM images from the literature, applied data curing and performed image analyses (fast Fourier transform, FFT; gray-level co-occurrence matrix, GLCM; histogram analysis, HA) and ML linear regression. The accuracy of the model did not considerably improve even by including AFM data in addition to the chemical structure fingerprints, material properties and process parameters. However, we found that a specific spatial wavelength of FFT (40-65 nm) significantly affects PCE. The GLCM and HA methods, such as homogeneity, correlation and skewness expand the scope of image analysis and artificial intelligence in materials science research fields.

Atomic Force Microscopy-Based Measurements of Retinal Microvessel Stiffness in Mice with Endothelial-Specific Deletion of CCN1.

Vascular stiffness is an independent predictor of human vascular diseases and is linked to ischemia, diabetes, high blood pressure, hyperlipidemia, and/or aging. Blood vessel stiffening increases owing to changes in the microscale architecture and/or content of extracellular, cytoskeletal, and nuclear matrix proteins. These alterations, while best appreciated in large blood vessels, also gradually occur in the microvasculature and play an important role in the initiation and progression of numerous microangiopathies including diabetic retinopathy. Although macroscopic measurements of arterial stiffness by pulse wave velocity are often used for clinical diagnosis, stiffness changes of intact microvessels and their causative factors have not been characterized. Herein, we describe the use of atomic force microscopy (AFM) to determine stiffness of mouse retinal capillaries and assess its regulation by the cellular communication network (CCN) 1, a stiffness-sensitive gene-encoded matricellular protein. AFM yields reproducible measurements of retinal capillary stiffness in lightly fixed freshly isolated retinal flat mounts. AFM measurements also show significant changes in compliance properties of the retinal microvasculature of mice with endothelial-specific deletion of CCN1, indicating that CCN1 expression, or lack thereof, affects the mechanical properties of microvascular cells in vivo. Thus, AFM has the force sensitivity and the spatial resolution necessary to measure the local modulus of retinal capillaries in situ and eventually to investigate microvascular compliance heterogeneities as key components of disease pathogenesis.

Nanomechanical properties of solid tumors as treatment monitoring biomarkers

Many tumors, such as types of sarcoma and breast cancer, stiffen as they grow in a host healthy tissue, while individual cancer cells are becoming softer. Tumor stiffening poses major pathophysiological barriers to the effective delivery of drugs and compromises treatment efficacy. It has been established that normalization of the mechanical properties of a tumor by targeting components of the tumor microenvironment (TME) enhances the delivery of anti-cancer agents and consequently the therapeutic outcome. Consequently, there is an urgent need for the development of biomarkers, which characterize the mechanical state of a particular tumor for the development of personalized treatments or for monitoring therapeutic strategies that target the TME. In this work, Atomic Force Microscopy (AFM) was used to assess human and murine nanomechanical properties from tumor biopsies. In the case of murine tumor models, the nanomechanical properties during tumor progression were measured and a TME normalization drug (tranilast) along with chemotherapy doxorubicin were employed in order to investigate whether AFM has the ability to capture changes in the nanomechanical properties of a tumor during treatment. The nanomechanical data were further correlated with ex vivo characterization of structural components of the TME. The results highlighted that nanomechanical properties alter during cancer progression and AFM measurements are sensitive enough to capture even small alterations during different types of treatments, namely normalization and chemotherapy. The identification of unique AFM-based nanomechanical properties can lead to the development of biomarkers for treatment prediction and monitoring. Statement of significanceCancer progression is associated with vast remodeling of the tumor microenvironment resulting in changes in the mechanical properties of the tissue. Indeed, many tumors stiffen as they grow and this stiffening compromises treatment efficacy. As a result, a number of treatments target tumor microenvironment in order to normalize its mechanical properties. Consequently, there is an urgent need for the development of innovative tools that can assess the mechanical properties of a particular tumor and monitor tumor progression and treatment outcomes. This work highlights the use of atomic force microscopy (AFM) for assessing the elasticity spectrum of solid tumors at different stages and during treatment. This knowledge is essential for the development of AFM-based nanomechanical biomarkers for treatment prediction and monitoring.

Modeling of carbon nanotubes (CNTs) and usage in drug delivery

One goal of device science is miniaturization; hence nanotechnology has received Considerable attention. The possibility that the unique properties of nanostructures will result in novel applications and devices is an enticing goal. Carbon nanotubes are allotropes of carbon with a cylindrical nanostructure. Nanotubes have been constructed with a length-to-diameter ratio of up to 28,000,000:1. Potential applications for carbon nanotubes (CNTs) include their use in atomic force microscopy and in adhesives, certainly chemical or mathematical investigation of nanotubes is very important but investigates both of them together has vital notability. there is considerable open literature related to the mathematical modeling of nanotubes, also many authors have investigated the chemical aspects of nanotubes. At the present, in this review, my aim is to both mathematical and chemical aspects of structured, characteristics are investigated.

Evaluation of the surface topography quality of large-area diamonds by image processing and mathematical modeling

Diamond with ultra-smooth and flat surface plays a crucial role in the various fields of nonlinear optics, NV center, waveguide, and so on, due to its remarkable physical properties. Consequently, the fast and efficient polishing and evaluating of diamond surfaces are indispensable to obtain high-quality smooth and flat diamond surfaces. As one of the most widespread techniques, atomic force microscope (AFM) and optical profilometry (OP) are enslaved to their small measurement regions and high time consuming, especially in the case of high-resolution measurement of large area diamond surfaces. Therefore, a novel approach to evaluate the polished diamond surface with high-efficiency and accuracy is desperately required. In this works, we propose a novel approach, surface topography quality (STQ) mathematical model, to achieve fast and large area evaluation to the polished diamond surface. Specifically, by combining currently popular image processing with mathematical statistics, STQ mathematical model generates a concept called surface topography quality rate (STQR) to quantitatively evaluate the surface quality of diamond. The results from large-area scanning electron microscope images before and after ion beam polishing demonstrates its reliability and preponderant advantage in dealing with large area surface compared to that of the conventional use of atomic force microscope. The mathematical model provides a unique and reliable approach to comprehensively and objectively evaluate diamond surface, which may promote the advancement of high-performance diamond-based devices.

Detection of Nanobubbles on Lubricant-Infused Surfaces Using AFM Meniscus Force Measurements.

So far, the presence of nanobubbles on lubricant-infused surfaces (LIS) has been overlooked, because of the difficulty in detecting them in such a complex system. We recently showed that anomalously large interfacial slip measured on LIS is explained by the presence of nanobubbles [Vega-Sánchez, Peppou-Chapman, Zhu and Neto, Nat. Commun., 2022 13, 351]. Crucial to drawing this conclusion was the use of atomic force microscopy (AFM) force-distance spectroscopy (meniscus force measurements) to directly image nanobubbles on LIS. This technique provided vital direct evidence of the spontaneous nucleation of nanobubbles on lubricant-infused hydrophobic surfaces. In this paper, we describe in detail the data collection and analysis of AFM meniscus force measurements on LIS and show how these powerful measurements can quantify both the thickness and distribution of multiple coexisting fluid layers (i.e., gas and oil) over a nanostructured surface. Using this technique, thousands of force curves were automatically analyzed. The results show that the interfacial tension of the nanobubbles is reduced from 52 ± 9 mN m-1 to 39 ± 4 mN m-1 by the presence of the silicone oil layer.

Photobiology of lipofuscin granules in the retinal pigment epithelium cells of the eye: norm, pathology, age.

Lipofuscin granules (LGs) are accumulated in the retinal pigment epithelium (RPE) cells. The progressive LG accumulation can somehow lead to pathology and accelerate the aging process. The review examines composition, spectral properties and photoactivity of LGs isolated from the human cadaver eyes. By use of atomic force microscopy and near-field microscopy, we have revealed the fluorescent heterogeneity of LGs. We have discovered the generation of reactive oxygen species by LGs, and found that LGs and melanolipofuscin granules are capable of photoinduced oxidation of lipids. It was shown that A2E, as the main fluorophore (bisretinoid) of LGs, is much less active as an oxidation photosensitizer than other fluorophores (bisretinoids) of LGs. Photooxidized products of bisretinoids pose a much greater danger to the cell than non-oxidized one. Our studies of the fluorescent properties of LGs and their fluorophores (bisretinoids) showed for the first time that their spectral characteristics change (shift to the short-wavelength region) in pathology and after exposure to ionizing radiation. By recording the fluorescence spectra and fluorescence decay kinetics of oxidized products of LG fluorophores, it is possible to improve the methods of early diagnosis of degenerative diseases. Lipofuscin ("aging pigment") is not an inert "slag". The photoactivity of LGs can pose a significant danger to the RPE cells. Fluorescence characteristics of LGs are a tool to detect early stages of degeneration in the retina and RPE.