The results of in-vivo two-photon imaging of lymphedema tissue are presented. The study involved 36 image samples from II stage lymphedema patients and 42 image samples from healthy volunteers. The papillary layer of the skin with a penetration depth of about 100 μm was examined. Both the collagen network disorganization and increase of the collagen/elastin ratio in lymphedema tissue, characterizing the severity of fibrosis, was observed. Various methods of image characterization, including edge detectors, a histogram of oriented gradients method, and a predictive model for diagnosis using machine learning, were used. The classification by “ensemble learning” provided 96% accuracy in validating the data from the testing set.

This paper addresses the application of multimodal nonlinear optical (MNLO) microscopy to clinical research within the context of label-free non-invasive molecular imaging. Here, a compact MNLO microscope based on a laser scanning microscope, a femtosecond laser, a time-correlated single-photon counting system, and a photonic crystal fiber are introduced for biomedical applications. By integrating two-photon fluorescence, two-photon fluorescence lifetime imaging, second-harmonic generation, and coherent anti-Stokes Raman scattering microscopy, the proposed scheme provides profound insights into the physicochemical properties related to 3D molecular orientation distribution, inter- and intra-molecular interactions, and disease progression in biological systems and organs. The high peak power and the low average intensity of near-infrared laser pulses allow for deep-penetration imaging without compromising sample vitality. Linking nonlinear optical phenomena with time/spectral/polarization-resolved imaging also makes it possible to obtain multidimensional information to address complex biomedical questions.

Despite the existing effective treatment methods, tuberculosis (TB) is the second most deadly infectious disease, its carriers in the latent and active phases accounting for more than 20% of the world population. An effective method for controlling TB and reducing TB mortality is regular population screening aimed at diagnosing the latent form of TB and taking preventive and curative measures. Numerous methods allow diagnosing TB by directly detecting Mycobacterium tuberculosis (M.tb) biomarkers, including M.tb DNA, proteins, and specific metabolites or antibodies produced by the host immune system in response to M.tb. PCR, ELISA, immunofluorescence and immunochemical analyses, flow cytometry, and other methods allow the detection of M.tb biomarkers or the host immune response to M.tb by recording the optical signal from fluorescent or colorimetric dyes that are components of the diagnostic systems. Current research in biosensors is aimed at increasing the sensitivity of detection, a promising approach being the use of fluorescent quantum dots as brighter and more photostable optical tags. Here, we review current methods for the detection of M.tb biomarkers using quantum dot-based nanosensors and summarize data on the M.tb biomarkers whose detection can be made considerably more sensitive by using these sensors.

The effect of low-dose photodynamic therapy on in vivo wound healing with topical application of 5-aminolevulinic acid and methylene blue was investigated using an animal model for two laser radiation doses (1 and 4 J/cm²). A second-harmonic-generation-to-auto-fluorescence aging index of the dermis (SAAID) was analyzed by two-photon microscopy. SAAID measured at 60–80 μm depths was shown to be a suitable quantitative parameter to monitor wound healing. A comparison of SAAID in healthy and wound tissues during phototherapy showed that both light doses were effective for wound healing; however, healing was better at a dose of 4 J/cm².

Bioinert materials such as the zirconium dioxide and aluminum oxide are widely used in surgery and dentistry due to the absence of cytotoxicity of the materials in relation to the surrounding cells of the body. However, little attention has been paid to the study of metabolic processes occurring at the implant–cell interface. The metabolic activity of mouse 3T3 fibroblasts incubated on yttrium-stabilized zirconium ceramics cured with aluminum oxide (ATZ) and stabilized zirconium ceramics (Y-TZP) was analyzed based on the ratio of the free/bound forms of cofactors NAD(P)H and FAD obtained using two-photon microscopy. The results show that fibroblasts incubated on ceramics demonstrate a shift towards the free form of NAD(P)H, which is observed during the glycolysis process, which, according to our assumptions, is related to the porosity of the surface of ceramic structures. Consequently, despite the high viability and good proliferation of fibroblasts assessed using an MTT test and a scanning electron microscope, the cells are in a state of hypoxia during incubation on ceramic structures. The FLIM results obtained in this work can be used as additional information for scientists who are interested in manufacturing osteoimplants.

The effect of low-dose photodynamic therapy on in vivo wound healing was investigated using optical coherence tomography. This work aims to develop an approach to quantitative assessment of the wound’s state during wound healing including the effect of low-dose photodynamic therapy using topical application of two different photosensitizers, 5-aminolevulinic acid and methylene blue, and two laser doses of 1 J/cm² and 4 J/cm². It was concluded that the laser dose of 4 J/cm² was better compared to 1 J/cm² and allowed the wound healing process to accelerate.