Search Results (85)

Background/Objectives: Motion artifacts significantly distort fluorescence measurements during surgical perfusion assessment, potentially leading to incorrect clinical decisions. This study evaluates the efficacy of automated motion compensation (MC) in quantitative indocyanine green (q-ICG) imaging to improve the accuracy of perfusion assessment. Methods: Frames from ICG perfusion assessment during 17 pancreaticoduodenectomies were analyzed. Regions of interest (ROIs) were systematically placed on each frame series, and automated MC was applied to track tissue movement. Performance was evaluated by comparing MC with surgeon-adjusted placement using multiple image quality metrics and analyzing perfusion metrics on time–intensity curves. Principal Component Analysis (PCA) was applied to explore whether image patterns could distinguish between successful and unsuccessful motion compensation. Results: Automated motion compensation successfully corrected motion artifacts in 67.5% of frame sequences, achieving comparable performance to surgeon-guided adjustments. PCA demonstrated clear separation between sufficient and insufficient corrections (AUC = 0.80). At the population level, MC did not significantly change perfusion slope (t(59) = 1.60, p = 0.11) or time-to-peak (Tmax; t(58) = 0.81, p = 0.42). Bland–Altman analysis showed a mean bias of −0.54 (SD = 3.32) for slope and 24.95 (SD = 238.40) for Tmax. At the individual level, 86.7% of slope and 79.7% of Tmax values differed by ≥10% after MC, with mean absolute percentage changes of 108.5% (median 37.8%) and 431.5% (median 65.9%), respectively. Conclusions: MC effectively reduces motion artifacts in fluorescence-guided perfusion assessment. By improving the precision of ICG-derived parameters, this technology enhances measurement reliability and represents an enabler for accurate intraoperative perfusion quantification. Full article

Indocyanine green near-infrared fluorescence (ICG-NIRF) imaging is widely used to assess tissue perfusion, yet its subjective interpretation limits correlation with postoperative parathyroid function. To address this, the Workflow model for ICG-angiography integrating Standardization and Quantification (WISQ) was developed. This exploratory prospective multicenter study evaluated the reproducibility of WISQ in adults undergoing total thyroidectomy at two Dutch university centres. Patients with contraindications to ICG or prior neck surgery were excluded. Intraoperative imaging used standardized camera settings with blood volume-adjusted ICG dosing, and perfusion curves were analyzed using predefined regions of interest. Eighty patients were included. Significant inter-centre variability was observed in maximum fluorescence intensity, inflow slope, and outflow slope (n = 30). At the lead centre, outflow was the most promising predictor of postoperative hypoparathyroidism (HPT) (median −0.33 [IQR −0.49–−0.15] a.f.u./s for HPT vs. −0.68 [−0.91–−0.41], n = 17, p = 0.08), although no parameter significantly predicted HPT. Repeated ICG injections consistently produced lower maximal intensities irrespective of injection rate, and reproducible curves were achieved only when ICG was freshly dissolved at 0.5 mg/mL instead of 2.5 mg/mL. These findings indicate that ICG concentration and injection technique influence perfusion kinetics and underscore the need to update WISQ with standardized injection dilution to improve its clinical utility. Full article

Nucleopeptides (NPs) are unnatural hybrid polymers designed by coupling nucleobases to the side chains of amino acid residues within peptides. In this study, we present the synthesis of an Fmoc-protected nucleobase amino acid (NBA) monomer (Fmoc-1,4-TzlNBA_A) with adenine attached to the side chain of L-homoazidoalanine (Aha) through a 1,4-linked-1,2,3-triazole. The coupling was accomplished by a Cu(I)-catalyzed azide–alkyne cycloaddition (CuAAC) of Fmoc-Aha and N9-propargyladenine. Subsequently, a homotrinucleopeptide (HalTzl_AAA) containing three 1,4-TzlNBA_A residues was synthesized, using different solid-phase peptide synthesis (SPPS) approaches, and its ability to recognize U-rich motifs of RNAs involved in the HIV replication cycle was studied using circular dichroism (CD) and fluorescence spectroscopy. CD curves confirmed the binding of HalTzl_AAA to U-rich motifs of the transactivation responsive element (TAR UUU RNA HIV-1) bulge and the anticodon stem–loop domain of human tRNALys3 (ASLLys3) by a decrease in the positive ellipticity band intensity around 265 nm during the complexation. 5′-(FAM(6))-labeled TAR UUU and hASLLys3 were used for fluorescence anisotropy binding studies. Fluorescence data revealed that HalTzl_AAA bound TAR’s UUU bulge with a moderate affinity (K_d ≈ 38 µM), whereas the ASLLys3 UUUU-containing loop sequence was recognized with 2.5 times lower affinity (with K_d ≈ 75 µM). Both the standard SPPS method and its variants, which involved the attachment of adenine to the L-Aha side chain using the click reaction during the synthesis on the resin or after the nucleopeptide cleavage, were characterized by a similar efficiency and yield. The CD and fluorescence results demonstrated that HalTzl_AAA recognized the U-rich sequences of the RNAs with moderate and varied affinities. It is likely that both the hydrogen bonds associated with the complementarity of the interacting sequences and the conformational aspects associated with the high conformational dynamics of U-rich motifs are important in the recognition process. The nucleopeptide represents a new class of RNA binders and may be a promising scaffold for the development of new antiviral drugs. Full article

The orange mullein is a biennial plant whose tall yellow flower spikes contain mucilage, saponins, and other medicinal compounds that have a beneficial effect on respiratory problems. As light quality is known to influence plant morphology and physiology, with effects often depending on the species, understanding these responses in mullein is of particular interest. Therefore, this study aimed to investigate the combined effects of different light-emitting diodes (white, red and blue) and their corresponding photon flux densities (PPFD) on the morphology, pigment composition, antioxidant activity, fluorescence parameters and OJIP transient curves in mullein (Verbascum phlomoides L.) seedlings. Seedlings grown under blue light, which had relatively higher PPFD, showed the greatest root length, leaf number, leaf and root fresh and dry biomass. Red light, with lower PPFD, resulted in the lowest values for these parameters. Compared to white light, pigment analysis showed that blue light increased chlorophyll a, total chlorophyll, carotenoid content, and the Chl a/b ratio. Also, blue light enhanced antioxidant activity, as well as the accumulation of phenolic compounds and flavonoids, indicating that it appeared to enhance the synthesis of secondary metabolites under this spectrum. In contrast, seedlings under red light exhibited the lowest ferric reducing antioxidant power values and tended to reduce levels of phenols and flavonoids, indicating a weaker antioxidative response. It was found that white light appeared to enhance the photochemical activity of photosystem II (PSII) and energy dissipation. Blue light improved linear electron transport, photosystem I (PSI) activity and overall photosynthetic performance. Red light preferentially increased electron flow towards the final acceptors of PSI, affecting the terminal part of the electron transport chain. Analysis of OJIP curves revealed spectrum and intensity-specific changes in the L, K, H, and G bands, demonstrating that light treatments with differing PPFDs selectively modulate PSII and PSI function. Full article

Background: Patients with chronic lymphocytic leukemia (CLL) who were not receiving treatment were included in this experimental prospective correlation study. We aimed to elucidate the complex relationship between smudge cells, surface CD20, and soluble CD20 in CLL patients. Methods: We created blood smears from blood samples collected from our patients using a manual technique consistently performed by the same technician. The May–Grunwald Giemsa dye was used to stain all of the slides. The B-cell phenotypic was analyzed using the FacsCanto II flow cytometer (Becton Dickinson, CA, USA) at the time of diagnosis. Competitive Enzyme-Linked Immunoassay (ELISA) was used to quantitatively assess the amounts of soluble CD20/MS4A1. Results: The percentage of smudge cells and soluble CD20 antigen levels were shown to be significantly inversely correlated, suggesting a considerable link (correlation coefficient (r) = −0.51, p = 0.006). Similarly, a significant inverse relationship (r = −0.36, p = 0.04) was found by the Spearman correlation test between the smudge cell ratio and CD20 median fluorescence intensity (MFI) on cell surfaces. Soluble CD20/MS4A1 and surface CD20 MFI were shown to have a weakly positive association that was almost statistically significant (Spearman’s rho = 0.34, p = 0.064). With a sensitivity of 69% and specificity of 86%, we discovered that a cut-off value of 2.2 ng/dL for soluble CD20 predicted higher smudge cells (area under the curve (95% confidence interval (CI)): 0.75 (0.57 to 0.93), p = 0.021). Conclusions: We found a significant inverse association between smudge cells and both surface CD20 and soluble CD20/MS4A1 in our study examining the correlation between smudge cells, soluble CD20, and CD20/MS4A1 in CLL patients. Our findings indicate that soluble CD20 may contribute to understanding the pathophysiology of smudge cells and could be further investigated as a potential prognostic marker in CLL. Full article

Harmful algal blooms (HABs) caused by toxic species such as Pseudo-nitzschia hasleana pose significant risks to marine ecosystems and human health. This study investigates the effects of heating rate on the fluorescence temperature curves (FTCs) of P. hasleana and compares them with non-toxic species (Phaeodactylum tricornutum and Picochlorum maculatum) to design a reliable detection method. An increasing heating rate leads to a change in the temperature spectrum of the fluorescence of the studied algae and to increasing differences between them. During the study, the FTCs were measured in the temperature range of 20–80 °C and at heating rates of 1, 2, 3, and 6°/min. The results showed that P. hasleana exhibited a distinct local fluorescence maximum at 45–55 °C when heated at a rate of 3 °C/min or more, which was absent in non-toxic species. Additionally, rapid heating (6 °C/min) preserved fluorescent pigment–protein complexes, yielding four-fold higher fluorescence intensity at 70–80 °C compared to slower rates. There were no such changes for the microalgae P. maculatum and P. tricornutum. The results of this study make it possible to increase the efficiency of detecting hazardous microalgae using non-invasive optical monitoring methods. These findings demonstrate that controlled heating protocols can enhance the species-specific identification of toxic microalgae, offering a practical tool for early HAB detection. Full article

Dental caries remains one of the most prevalent global diseases, and the early detection of occlusal lesions is critical because demineralization often begins deep within pits and fissures where conventional visual–tactile or radiographic inspection cannot detect it. SmarTooth, a newly introduced fluorescence device that irradiates enamel with a 655 nm laser and records the reflected intensity, may provide more objective, quantitative diagnoses. This study assessed its diagnostic performance against the International Caries Detection and Assessment System (ICDAS). We examined 1421 occlusal surfaces from 153 adults, scored each surface with ICDAS codes 0–4, and recorded SmarTooth peak values. Spearman’s rank correlation was used to test the association between codes and peak values; one-way ANOVA with Tukey’s post hoc was used to compare mean values across codes; and sensitivity, specificity, and the area under the receiver operating characteristic curve (AUROC) were calculated at three diagnostic thresholds: D1 (0 vs. 1–4), D2 (0–2 vs. 3–4), and D3 (0–3 vs. 4). The SmarTooth values rose with lesion severity and correlated moderately with ICDAS (r = 0.495, p < 0.001). The AUROC ranged from 0.69 to 0.82, with the best accuracy observed at D2 (cut-off: 7.0; AUC: 0.82; sensitivity: 78.3%; specificity: 77.4%). These findings suggest that SmarTooth can complement ICDAS scoring as an adjunctive tool, potentially enhancing diagnostic accuracy and supporting early intervention for occlusal caries in general dental practice. Full article

This study presents a biomonitoring study of surface waters in the Krynka River basin, encompassing three major regional reservoirs: Khanzhenkovskoe, Olkhovskoe, and Zuyevskoe. These water bodies face significant anthropogenic pressure from mining effluents, industrial discharges, and domestic wastewater. Key pollutants identified are surfactants (SAAs), sulfates, phenols, chlorides, and manganese, with chemical oxygen demand (COD) exceeding regulatory limits. The research was conducted in September 2024. Based on the Specific Combinatorial Water Pollution Index, surface waters in the studied objects can be characterized as slightly polluted. To assess the negative impact of the identified pollutants on hydrobionts, the species composition of phytoplankton of the studied water bodies was analyzed. In the Olkhovskoe Reservoir and Olkhovaya River, cyanobacterial blooms (Oscillatoria agardhii G.) were observed, altering biodiversity in the Krynka River and Zuyevskoe Reservoir. Phytoplankton genera Synedra, Amphiprora, and Navicula—established bioindicators of aquatic ecosystem health—were dominant in Khanzhenkovskoe Reservoir, signaling nutrient enrichment and organic pollution. Changes in the species composition and structure of phytoplankton in the Krynka River, its tributaries and reservoirs, indicate a change in the level of saprobic water bodies from β to α-mesosaprobic, which indicates both the general level of surface water pollution and the accumulation of pollutants along the course of the river. The paper presents the results of fluorimetric analysis of photosynthetic activity of natural phytoplankton cells and demonstrates the possibility of using fluorescence induction curves for regular monitoring measurements. Fluorescence parameters indicate a general deterioration of photosynthetic activity of natural phytoplankton. The growth of Oscillatoria agardhii in the waters of the Olkhovskoe Reservoir and of green microalgae in the Zuevskoe Reservoir led to an increase in the fluorescence quantum yield (F_v/F_m) and the total photosynthetic activity index (PI), which makes it possible to use these parameters as indicator parameters reflecting the intensity of “blooming” of various phytoplankton species. Full article

Goldfish (Carassius auratus) gills function as both respiratory and immune-regulatory organs, integrating neuroendocrine and immune responses to environmental stimuli. This study explores the spatial organization and interaction of neuroendocrine cells (NECs) and immune cells within goldfish gills using confocal immunohistochemistry and transmission electron microscopy. NECs, identified near blood capillaries and nerve fibers, highlight their role in environmental sensing and physiological regulation. These cells express serotonin (5-HT), a neurotransmitter critical for neuroimmune communication. Two distinct macrophage subsets were observed: iNOS-positive macrophages, concentrated in the basal epithelium, suggest a pro-inflammatory role, whereas 5-HT-positive macrophages, dispersed in the subepithelium, likely contribute to immune modulation. The co-localization of MHC-II and CD68 in macrophages further supports an active antigen-processing system in the gills. Ultrastructural analysis revealed diverse immune cells, including rodlet cells, telocytes, and lymphocytes, within the gill epithelium. Telocytes formed intricate networks with immune cells, highlighting their role in immune coordination and tissue homeostasis. These findings provide new insights into the neuroimmune interactions in fish gills, contributing to a broader understanding of aquatic immune systems and environmental adaptability. Full article

In recent years, rare earth ion and transition metal ion co-doped fluorescent materials have attracted a lot of attention in the fields of WLEDs and optical temperature sensing. In this study, I successfully prepared the dual-emission Mg₃Ga₂SnO₈:Eu³⁺,Mn⁴⁺ red phosphors and the XRD patterns and refinement results show that the prepared phosphors belong to the Fd-3m space group. The energy transfer process between Eu³⁺ and Mn⁴⁺ was systematically investigated by emission spectra and decay curves of Mg₃Ga₂SnO₈:0.12Eu³⁺,yMn⁴⁺ (0.002 ≤ y ≤ 0.012) phosphors and the maximum value of transfer efficiency can reach 71.2%. Due to the weak thermal quenching effect of Eu³⁺, its emission provides a stable reference for the rapid thermal quenching of the Mn⁴⁺ emission peak, thereby achieving good temperature measurement performance. The relative thermometric sensitivities of the fluorescence intensity ratio and fluorescence lifetime methods reached a maximum value of 2.53% K⁻¹ at 448 K and a maximum value of 3.38% K⁻¹ at 473 K. In addition, the prepared WLEDs utilizing Mg₃Ga₂SnO₈:0.12Eu³⁺ phosphor have a high color rendering index of 82.5 and correlated color temperature of 6170 K. The electroluminescence spectrum of the synthesized red LED device by Mg₃Ga₂SnO₈:0.009Mn⁴⁺ phosphor highly overlaps with the absorption range of the phytochrome P_FR and thus can effectively promote plant growth. Therefore, the Mg₃Ga₂SnO₈:Eu³⁺,Mn⁴⁺ phosphors have good application prospects in WLEDs, temperature sensing, and plant growth illumination. Full article

Voltage-gated sodium (Nav) channels play a crucial role in initiating and propagating action potentials throughout the heart, muscles and nervous systems, making them targets for a number of drugs and toxins. While patch-clamp electrophysiology is considered the gold standard for measuring ion channel activity, its labor-intensive and time-consuming nature highlights the need for fast screening strategies to facilitate a preliminary selection of potential drugs or hazards. In this study, a high-throughput and cost-effective biosensing method was developed to rapidly identify specific agonists and inhibitors targeting the human Nav1.1 (hNav1.1) channel. It combines a red fluorescent dye sensitive to transmembrane potentials with CHO cells stably expressing the hNav1.1 α-subunit (hNav1.1-CHO). In the initial screening mode, the tested compounds were mixed with pre-equilibrated hNav1.1-CHO cells and dye to detect potential agonist effects via fluorescence enhancement. In cases where no fluorescence enhancement was observed, the addition of a known agonist veratridine allowed the indication of inhibitor candidates by fluorescence reduction, relative to the veratridine control without test compounds. Potential agonists or inhibitors identified in the initial screening were further evaluated by measuring concentration–response curves to determine EC₅₀/IC₅₀ values, providing semi-quantitative estimates of their binding strength to hNav1.1. This robust, high-throughput biosensing assay was validated through comparisons with the patch-clamp results and tested with 12 marine toxins, yielding consistent results. It holds promise as a low-cost, rapid, and long-term stable approach for drug discovery and non-target screening of neurotoxins. Full article

The tomato is among the crops with the most extensive cultivated area and greatest consumption in our nation; nonetheless, secondary salinization of facility soil significantly hinders the sustainable growth of facility agriculture. Melatonin (MT), as an innovative plant growth regulator, is essential in stress responses. This research used a hydroponic setup to replicate saline stress conditions. Different endogenous levels of melatonin (MT) were established by foliar spraying of 100 μmol·L⁻¹ MT, the MT synthesis inhibitor p-CPA (100 μmol·L⁻¹), and a combination of p-CPA and MT, to investigate the mechanism by which MT mitigates the effects of salt stress on the photosynthetic efficiency of tomato seedlings. Results indicated that after six days of salt stress, the endogenous MT content in tomato seedlings drastically decreased, with declines in the net photosynthetic rate and photosystem performance indices (PI_total and PI_abs). The OJIP fluorescence curve exhibited distortion, characterized by anomalous K-band and L-band manifestations. Exogenous MT dramatically enhanced the gene (TrpDC, T5H, SNAcT, and AcSNMT) expression of critical enzymes in MT synthesis, therefore boosting the level of endogenous MT. The application of MT enhanced the photosynthetic parameters. MT treatment decreased the fluorescence intensities of the J-phase and I-phase in the OJIP curve under salt stress, attenuated the irregularities in the K-band and L-band performance, and concurrently enhanced quantum yield and energy partitioning ratios. It specifically elevated φP_o, φE_o, and ψ_o, while decreasing φD_o. The therapy enhanced parameters of both the membrane model (ABS/RC, DI_o/RC, ET_o/RC, and TR_o/RC) and leaf model (ABS/CS_m, TR_o/CS_m, ET_o/CS_m, and DI_o/CS_m). Conversely, the injection of exogenous p-CPA exacerbated salt stress-related damage to the photosystem of tomato seedlings and diminished the beneficial effects of MT. The findings suggest that exogenous MT mitigates salt stress-induced photoinhibition by (1) modulating endogenous MT concentrations, (2) augmenting PSII reaction center functionality, (3) safeguarding the oxygen-evolving complex (OEC), (4) reinstating PSI redox potential, (5) facilitating photosynthetic electron transport, and (6) optimizing energy absorption and dissipation. As a result, MT markedly enhanced photochemical performance and facilitated development and salt stress resilience in tomato seedlings. Full article

Background/Objectives: Potency testing of clinical-grade lentiviral vectors (LVVs) is critical to support a drug’s commercial approval. Careful consideration should be paid to the development of a suitable potency test during the drug’s clinical development. We aimed to develop an affordable, quantitative test for our CAR19-LVV, based on a measure of transgene’s functional activity. Methods: Several indicators of functional activity of CAR19-LVV were explored in a co-culture setting of CAR-transduced Jurkat cells and CD19-expressing target cells. The selected assay was further developed and subjected to validation. Assay’s adaptability to other CAR-encoding LVV and autologous CAR-T cell products was also investigated. Results: Measure of CD69 expression on the membrane of Jurkat-CAR-expressing cells is a specific indicator of CAR functionality. Quantification of CD69 in terms of mean fluorescence intensity (MFI), coupled with an intra-assay standard curve calibration, allows for a quantitative assay with high precision, specificity, robustness, linearity and accuracy. The assay has also shown optimal performance for a CARBCMA-LVV product. Importantly, we show that in primary T cells, CD69 expression reflects CAR-T cell cytotoxicity. After adaptation, we have applied a CD69-based potency test, with simultaneous measurement of CAR-T cell cytotoxicity, to autologous CAR-T cell products, demonstrating the assay’s specificity also in this context. Conclusions: We developed a validated, in vitro cell-based potency test, using a quantitative flow-cytometry method, for our CAR19-LVV. The assay is based on the detection of T-cell activation upon CAR binding to antigen, which is a measure of transgene functionality. The assay was easily adapted to another CAR-encoding LVV, targeting a different molecule. Furthermore, the same assay principle can be applied in the context of autologous CAR-T cell products. The quantitative CD69 potency assay shows reduced variability among autologous products compared to the IFNγ assay and allows for simultaneous evaluation of traditional semi-quantitative cytotoxicity, thereby directly evaluating the drug’s mechanism of action (MoA) in the same assay. Full article

Point-of-care (POC) testing has revolutionized diagnostics by providing rapid, accessible solutions outside traditional laboratory settings. However, many POC systems lack the sensitivity or multiplexing capability required for complex diseases. This study introduces an LED-based fluorescence reader designed for POC applications, enabling multiplex detection of lupus nephritis (LN) biomarkers using a biomarker microarray (BMA) slide. The reader integrates an LED excitation source, neutral density (ND) filters for precise intensity control, and onboard image processing with Gaussian smoothing and centroid thresholding to enhance signal detection and localization. Five LN biomarkers (VSIG4, OPN, VCAM1, ALCAM, and TNFRSF1B) were assessed, and performance was validated against a Genepix laser-based scanner. The LED reader demonstrated strong correlation coefficients (r = 0.96–0.98) with the Genepix system for both standard curves and patient samples, achieving robust signal-to-noise ratios and reproducibility across all biomarkers. The multiplex format reduced sample volume and allowed simultaneous analysis of multiple biomarkers. These results highlight the reader’s potential to bridge the gap between laboratory-grade precision and POC accessibility. By combining portability, cost-effectiveness, and high analytical performance, this fluorescence reader provides a practical solution for POC diagnostics, particularly in resource-limited settings, improving the feasibility of routine monitoring and early intervention for diseases requiring comprehensive biomarker analysis. Full article

Chemical weed control is a significant agricultural concern, and reliance on a limited range of herbicide action modes has increased resistant weed species, many of which use C4 metabolism. As a result, the identification of novel herbicidal agents with low toxicity targeting C4 plants becomes imperative. An assessment was conducted on the impact of 3-cyanobenzoic acid on the growth and photosynthetic processes of maize (Zea mays), a representative C4 plant, cultivated hydroponically over 14 days. The results showed a significant reduction in plant growth and notable disruptions in gas exchange and chlorophyll a fluorescence due to the application of 3-cyanobenzoic acid, indicating compromised photosynthetic activity. Parameters such as the chlorophyll index, net assimilation (A), stomatal conductance (g_s), intercellular CO₂ concentration (C_i), maximum effective photochemical efficiency (F_v′/F_m′), photochemical quenching coefficient (q_P), quantum yield of photosystem II photochemistry (ϕ_PSII), and electron transport rate through PSII (ETR) all decreased. The A/PAR curve revealed reductions in the maximum net assimilation rate (A_max) and apparent quantum yield (ϕ), alongside an increased light compensation point (LCP). Moreover, 3-cyanobenzoic acid significantly decreased the carboxylation rates of RuBisCo (V_cmax) and PEPCase (V_pmax), electron transport rate (J), and mesophilic conductance (g_m). Overall, 3-cyanobenzoic acid induced substantial changes in plant growth, carboxylative processes, and photochemical activities. The treated plants also exhibited heightened susceptibility to intense light conditions, indicating a significant and potentially adverse impact on their physiological functions. These findings suggest that 3-cyanobenzoic acid or its analogs could be promising for future research targeting photosynthesis. Full article