New Aspects Regarding the Fluorescence Spectra of Melanin and Neuromelanin in Pigmented Human Tissue Concerning Hypoxia.

Melanin is a crucial pigment in melanomagenesis. Its fluorescence in human tissue is exceedingly weak but can be detected through advanced laser spectroscopy techniques. The spectral profile of melanin fluorescence distinctively varies among melanocytes, nevomelanocytes, and melanoma cells, with melanoma cells exhibiting a notably "red" fluorescence spectrum. This characteristic enables the diagnosis of melanoma both in vivo and in histological samples. Neuromelanin, a brain pigment akin to melanin, shares similar fluorescence properties. Its fluorescence can also be quantified with high spectral resolution using the same laser spectroscopic methods. Documented fluorescence spectra of neuromelanin in histological samples from the substantia nigra substantiate these findings. Our research reveals that the spectral behavior of neuromelanin fluorescence mirrors that of melanin in melanomas. This indicates that the typical red fluorescence is likely influenced by the microenvironment around (neuro)melanin, rather than by direct pigment interactions. Our ongoing studies aim to further explore this distinctive "red" fluorescence. We have observed this red fluorescence spectrum in post-mortem measurements of melanin in benign nevus. The characteristic red spectrum is also evident here (unlike the benign nevus in vivo), suggesting that hypoxia may contribute to this phenomenon. Given the central role of hypoxia in both melanoma development and treatment, as well as in fundamental Parkinson's disease mechanisms, this study discusses strategies aimed at reinforcing the hypothesis that red fluorescence from (neuro)melanin serves as an indicator of hypoxia.

Cross-section measurements of multilayer automotive paint samples using combined Raman spectroscopy and LIBS.

Multilayer automotive paint samples can be important evidence in forensic investigations but due to their inherent complexity it is a challenging task to analyze them. We herein present a method for a comprehensive chemical sample characterization based on the subsequent measurements of paint samples with Raman spectroscopy and laser induced breakdown spectroscopy (LIBS) at the same sampling points along the cross-sections of the paints. The method requires minimal sample preparation because the layers do not have to be separated. In combination with multivariate data analysis (namely k-means) the proposed method provides information both on the distribution of molecular compounds as well as elements which could then be used for the discrimination of different paints. Eight different samples were investigated and individual chemical profiles were found whereby a spatial resolution of close to 10 µm could be demonstrated. Investigation of different samples of the same origin showed similar profiles revealing the reproducibility of the proposed method. We therefore believe that our approach could be a good addition to the methods used so far for the analysis of paint samples.

From Melanocytes to Melanoma Cells: Characterization of the Malignant Transformation by Four Distinctly Different Melanin Fluorescence Spectra (Review).

The melanin fluorescence emitted by pigment cells of the human skin has been a central research topic for decades, because melanin, on the one hand, protects against (solar) radiation in the near-UV range, whereas on the other hand, melanocytes are the starting point for the malignant transformation into melanoma. Until recently, however, melanin fluorescence was not accessible in the context of conventional spectroscopy, because it is ultraweak and is overshadowed by the more intense so-called autofluorescence of endogenous fluorophores. The advent of a new method of laser spectroscopy has made this melanin fluorescence measurable in vivo. A stepwise two-photon absorption with 800 nm photons is used, which more selectively excites melanin (dermatofluoroscopy). Our review summarizes the experimental results on melanin fluorescence of the four types of cutaneous pigment cells from healthy and malignant tissues. Outstanding is the finding that different types of melanocytes (i.e., melanocytes of common nevi, versus dysplastic nevi or versus melanoma cells) show characteristically different fluorescence spectra. The possibilities of using this melanin fluorescence for melanoma diagnosis are shown. Moreover, the uniform fluorescence spectra emitted by different melanoma subtypes are essential. Conclusions are drawn about the molecular processes in the melanosomes that determine fluorescence. Finally, experimental suggestions for further investigations are given.

A phenomenological and quantitative view on the degradation of positive electrodes from spent lithium-ion batteries in humid atmosphere.

The present study deals with the phenomenological observation of the corrosion of the positive electrode foil of lithium-ion batteries containing LiNi0.6Co0.2Mn0.2O2 (NMC) as cathode material. Due to the presence of moisture, localized water accumulation is formed on the NMC surface. The water absorbed by the electrolyte reacts with the NMC under Li+/H+ exchange and the resulting pH increase leads to dissolution of the carrier foil and characteristic salt-like blooms on the NMC surface. With the increase in the relative area occupied by the holes in the aluminum foil per time, a sufficiently suitable parameter was found with which to quantitatively determine the extent of corrosion. The degree of degradation depends on time and ambient humidity. It was shown that functional recycling with the water jet method is no longer applicable for degraded foils, since the mechanical stability of the foils decreases as corrosion progresses. Lithium, aluminum, sulfur and oxygen were detected in the blooms using SEM-EDX and Laser-Induced-Breakdown-Spectroscopy (LIBS). The underlying NMC layer was found to contain mainly aluminum and significantly lower lithium content than the non-degraded material. SEM and Raman microscopy analyses also showed that the active material is also locally degraded and therefore no longer suitable for functional recycling.

Criteria for the multiple use of isolated perfused hearts in electrophysiological and metabolic experiments.

Both ethical and economic restrictions limit the availability of porcine hearts for in vitro perfusion experiments. Therefore, we tested the feasibility of multiple use of in vitro perfused working hearts for electrophysiological and metabolic investigations. Pig hearts (n=7) rejected for originally planned haemodynamic measurements because of exclusion criteria were perfused in a four-chamber working heart mode. All hearts were kept in steady-state conditions on a low haemodynamic level over 2 h during 75-channel ECG recordings and NADH fluorescence measurements before and after norepinephrine (NE) was administered. QRS and QT interval durations were in a range comparable to in vitro studies and, like QRS and T amplitudes, were found to be sensitive markers of the changing condition of the isolated heart preparation, as myocardial oedema leads to prolonged QRS and QT intervals and declining ECG voltage amplitudes. A change in NADH fluorescence following NE administration was observed in the first 150 min of perfusion, but not later. Considering a time frame of 120 min, multiple use of isolated perfused porcine hearts with low-level haemodynamics may allow a broad spectrum of investigations and could therefore represent a possibility of overcoming the restricted availability of porcine hearts.

Fluorimetric characterisation of metabolic activity of ex vivo perfused pig hearts.

Autofluorescence of tissues and organs is an indicator of the physiological state of cells. The aim of the study was to investigate whether fluorimetric determination of the redox state of the ex vivo perfused pig heart can provide fast online detection of progressive changes in heart muscle tissue. Measurements on six organs perfused in a four-chamber working heart model were performed using a spectroscopic method exploiting the specific and different fluorescence lifetimes of intrinsic fluorophores such as NADH and flavins and providing a means of internal signal referencing. It was shown that the redox potential of heart muscle tissue can be assessed by fluorescence measurement. In the steady-state phase of the beating heart, spectroscopic measurements revealed a change in redox state from an initial constant level to a continuous decrease, accompanied by a decrease in heart performance and indications of changes in electrolyte equilibrium (K(+) concentration). At the same time, troponin I levels in the perfusate increased. The results indicate that fluorimetric determination of heart muscle metabolic activity yields reliable information about the functional status of the ex vivo heart and may be advantageous for the optimisation of ex vivo organ models.

Screening scheme based on measurement of fluorescence lifetime in the nanosecond domain.

The authors demonstrate that the fluorescence lifetime of certain fluorescent labels is a useful parameter to detect affinity binding between biotin and streptavidin, as well as between biotinylated bovine serum albumin and streptavidin. The assay is performed in a microplate format, and lifetimes are determined using dye laser-induced fluorescence. Four fluorescent labels are presented that undergo a significant change in their lifetime upon affinity binding. The scheme, referred to as the fluorescence lifetime affinity assay, has several attractive features in that it requires single labeling only, represents a homogeneous assay, allows each of the 2 binding partners to be labeled, and is compatible with the standard microwell formats used in high-throughput screening.

UV-induced fluorescence spectra and lifetime determination for detection of leaf rust (Puccinia triticina) in susceptible and resistant wheat (Triticum aestivum) cultivars.

In modern agriculture, the use of cultivars that are resistant against specific stresses, e.g. pathogen infections, is an integral component. Considering the great demand for a rapid and objective screening method for stress resistance of new cultivars, the question arises, whether time resolved fluorescence spectroscopy is suitable for such purposes. Amongst others, infected plants might accumulate specific compounds such as salicylic acid and phenylpropanoid compounds as key substances in plant disease resistance, whereas synthesis and accumulation may influence fluorescence parameters such as absolute intensity of single peaks, ratios between peaks and lifetime. Experiments were conducted in a controlled-environment cabinet cultivating four leaf rust susceptible and three leaf rust resistant genotypes. Fluorescence measurements were conducted using a compact fibre-optic fluorescence spectrometer with a nanosecond time-resolution. Results of experiments revealed that UV-induced measurements of spectral characteristics as well as determination of fluorescence lifetime are suited to detect leaf rust (Puccinia triticina) in wheat (Triticum aestivum L.) cultivars as early as 2 days after inoculation (dai). For this purpose several parameters such as the fluorescence (F) amplitude ratios F451/F522, F451/F687, F451/F736, F522/F687, F522/F736 as well as fluorescence mean lifetime especially at 470nm, might be used. Discrimination between resistant and susceptible cultivars to the leaf rust pathogen could be accomplished 3dai by using the ratio of fluorescence amplitude between the blue (F451nm) and red (F687nm) peak, and mean lifetime at 440, 500 and 530nm. Our results indicate that the combination of spectrally and time-resolved fluorescence could be an additional tool in plant breeding programs for an automatic and precise high-throughput system for evaluation of the pathogen resistance of new genotypes.

Improved routine bio-medical and bio-analytical online fluorescence measurements using fluorescence lifetime resolution.

Fluorescence techniques are widely used as sensitive detection methods in bio-analytics. The use of the bio-physical parameter fluorescence lifetime additional to the spectral characteristics of fluorescence has the potential to improve fluorescence-related detection methods in terms of selectivity in signal recognition, robustness against disturbing influences, and the accessibility of novel bio-chemical process parameters. This article describes the technical set up of a time-resolving instrument with either a fixed time-gated detection principle for improved evaluation of tissue metabolism by an online monitoring of the tissue autofluorescence or a direct fluorescence lifetime detection principle for lifetime-based fluorescent assays.