Characterization of a 1,4-dithiane gold self-assembled monolayer: an electrochemical sensor for the cyt-c redox process

Electrochemical desorption and spectroscopic investigations of the gold electrode surface modified with 1,4-dithiane (1,4-dt) organothiol species were performed. The wave observed at ?0.87 V versus Ag ∣ AgCl in the LSV (linear sweep voltammetry) reductive curve of the 1,4-dt compared to that for a similar 4-mercaptopyridine (pyS) system (?0.56 V) is indicative of a most effectively chemisorbed monolayer. The evaluation of the capability of the 1,4-dt self-assembled monolayer (SAM) in assessing the direct electron transfer (ET) of cytochrome c (cyt c) metalloprotein was investigated by cyclic voltammetry. The electrochemical response of the cyt c (E_1/2 ≈0.0 V vs. Ag ∣ AgCl, ΔE_p ≈50 mV) showed the characteristics of a reversible redox process. The cyt c voltammetric parameters acquired with the 24-h air exposure modified electrode, and after 100 cycles suggest a considerable improvement of the 1,4-dt electrode performance. The surface enhanced Raman spectroscopy (SERS) spectra revealed that 1,4-dt species is in a mixed gauche and trans orientation on the gold surface. The shift for higher wavenumbers observed for the C–S stretching modes in the SERS spectra, comparatively to the normal Raman spectrum, is assigned to the 1,4-dt coordination to surface gold atoms via a π interaction with the sulfur p-orbitals. The data collected suggest that this π interaction plays an important role on the stability of the 1,4-dt adlayer, improving the assessment of the cyt c heterogeneous electron transfer reaction.     

Study of redox mechanism of poly(o-aminophenol) using in situ techniques: evidence of two redox processes

The poly(o-aminophenol) (POAP) redox process has been studied in aqueous acid solution using spectroscopic and optic in situ techniques. The redox transition of the POAP from its completely oxidized state to its completely reduced state occurs through two consecutive reactions in which a charged intermediate species takes part. UV–Vis and Raman signals agree with an increase of the concentration of an intermediate species until the potential of the maximum redox peak, which later diminishes with the potential. The results of in situ FTIR spectroscopy agrees with Raman measurements. Probe beam deflection (PBD) suggests that during its oxidation, the polymer incorporates anions in a first process and then expels protons in a second one.     

Electrosynthesis and characterisation of poly(N-methylaniline) in organic solvents

The electropolymerisation of N-methylaniline was studied on glassy carbon and optically transparent tin oxide electrodes in the organic solvents dimethylformamide (DMF) and dimethyl sulfoxide (DMSO) containing 1.0 M methanesulfonic acid or 1.0 M trifluoromethanesulfonic acid. Our results show that a thin film of poly(N-methylaniline) can be obtained in DMF and DMSO, although the reaction product is very soluble. The PNMA film and the soluble fraction of the reaction product was analyzed and characterised with cyclic voltammetry, mass spectroscopy, in situ UV–Visible and Raman spectroscopy. The formation of an intermediate in DMF with an absorbance maximum at 720 nm was confirmed with in situ UV–Visible measurements. This is in contrast to polymerisation in aqueous solution where the absorbance maximum of the intermediate has been reported to appear at 441–460 nm.     

Characterizing Electric Field Exposed P3HT Thin Films Using Polarized‐Light Spectroscopies

P3HT (poly (3‐hexylthiophene)) has been widely used as a donor in the active layer in organic photovoltaic devices. Although moderately high‐power conversion efficiencies have been achieved with P3HT‐based devices, structural details, such as the orientation of polymer units and the extent of H‐ and J‐aggregation are not yet fully understood; and different measures have been taken to control the ordering in the material. One such measure, which has been exploited, is to apply an electric field from a Van de Graaff generator. Fluorescence (to measure anisotropy instead of polarization, which is more commonly measured) and Raman spectroscopy are used to characterize the order of P3HT molecules in thin films resulting from the field. Preferential orientations of the units in a thin film are determined, consistent with observed hole mobility in thin‐film transistors, and it is observed that the apparent H‐coupling strength changes when the films are exposed to oriented electrical fields during drying.

     

拉曼光谱研究两面针活性成分诱导肝癌细胞的凋亡

运用拉曼光谱法对两面针活性成分诱导的肝癌细胞凋亡进行分析,肝癌细胞7404分别经10 mg·L~(-1)氯化两面针碱及3 g·L~(-1)两面针提取液处理后,收集经药液处理12,24,36,48 h的各组细胞的拉曼光谱后,通过Hochest33342/PI荧光染色法鉴定细胞并保留荧光染色阳性(发生凋亡活细胞)的光谱,并在Origin Pro8.0系统中比较各组平均光谱的差异。收集肝癌细胞的拉曼光谱依次进行背景扣除、平滑、归一化等方法处理。Hochest荧光染色后空白组细胞核染色均匀,而药物处理48 h后,核碎裂,拉曼光谱结果显示两面针提取液处理肝癌细胞12,24,36,48 h后,与核酸及蛋白质相关的峰均有降低,其中785,1 002,1 175,1 660 cm~(-1)峰强度随两面针药物作用时间的延长而降低,表明两面针活性成分能够诱导肝癌细胞凋亡,凋亡的肝癌细胞中核酸和蛋白质的含量均低于活细胞。两面针活性成分作用时间与药效呈一定的相关性。拉曼光谱能够反映中药两面针活性成分作用后的肝癌细胞内物质变化的信息,对实时监测细胞凋亡过程及药物的临床应用具有重要意义。     

Glass-ionomer and calcium silicate-based cements interactions with human dentine in health and disease: Two-photon fluorescence microscopy and Raman spectroscopy analysis.

ObjectivesTo investigate the potential mineralising effects of calcium silicate-based dentine replacement material (Biodentine?) in comparison with glass-ionomer cement (GIC) (Fuji IX?) on different human dentine substrates using a multimodal non-invasive optical assessment.MethodsCements were applied on artificially demineralised or naturally carious dentine and stored for 4 weeks in phosphate-rich media +/- tetracycline used for mineralisation labelling. Interfacial dentine was examined from the same sample and location before and after aging using two-photon fluorescence microscopy, fluorescence lifetime imaging (FLIM) and second harmonic generation (SHG) imaging. Additionally, Raman spectroscopy was used to detect changes in the mineral content of dentine.ResultsSignificant changes in the fluorescence intensity and lifetime were detected in partially demineralised dentine and caries-affected dentine underneath both tested cements, after storage (p < 0.001). This was associated with a significant increase in the mineral content as indicated by the increased intensity of the phosphate Raman peak located at 959 cm^?1 (p < 0.0001). Caries-infected dentine showed significant fluorescence changes under Biodentine? after storage (p < 0.001), but not under GIC (p = 0.44). Tetracycline binding induced a reduction in the fluorescence lifetime with comparable increase in the fluorescence intensity in both cements’ groups within the affected dentine (p < 0.001). SignificanceTwo-photon fluorescence microscopy can be used efficiently for non-destructive in-vitro dentine caries characterisation providing a technique for studying the same dentine-cement interface over time and detect changes. Biodentine? demonstrated comparable remineralising potential to GIC, in addition to inducing remineralisation of caries-infected dentine. This may suggest using Biodentine? as part of minimally invasive operative dentistry (MID) in caries management.     

基于重水拉曼技术的氯己定对白色念珠菌 抑菌效能的研究

目的评价重水拉曼技术的普适性及氯己定(CHX)对白色念珠菌的抑菌效能。方法1)采用分光光度计测定吸光度值OD600,重水拉曼技术测定C?D ratio(重水峰所占比例)随时间的变化,探索重水对白色念珠菌生长的影响及该菌对重水的吸收规律,以评价重水拉曼技术的普适性。2)采用肉汤稀释法和重水拉曼技术,测定CHX对白色念珠菌的最低抑菌浓度(MIC)及最小代谢活性抑制浓度(MIC?MA),评价CHX对白色念珠菌生长及代谢的抑制效果。结果1)白色念珠菌在质量分数≤30%重水中,生长未受到明显抑制(P>0.05);白色念珠菌能活跃代谢重水并通过拉曼图谱检测,且C?D ratio与重水质量分数呈线性正相关关系(R^2=0.9894,P<0.05)。2)CHX对白色念珠菌的MIC为4μg·mL^-1,MIC?MA为8μg·mL^-1。在MIC下,白色念珠菌的生长受到完全抑制,但仍具有很高的代谢活性;只有达到2×MIC,即MIC?MA,该菌的代谢才能得到完全抑制。结论重水拉曼技术适用于评价药物对真菌代谢活性的影响,临床常用质量分数的CHX可完全抑制白色念珠菌的生长及代谢。     

Optical spectroscopy and imaging for early lung cancer detection: a review

Over the last 15 years, optical spectroscopy and imaging has been intensively studied to improve the detection and localization of early lung cancer. Autofluorescence bronchoscopy (AFB) is the most successfully developed technique and has significantly improved the detection sensitivity of early lung cancer. In this review, the optical principles behind white-light and autofluorescence bronchoscopy, as well as the role of AFB in the diagnosis of early lung cancer and the overall management of patients with early lung cancer are discussed. Other newest development such as Raman spectroscopy and simultaneous imaging and spectroscopy measurements are also highlighted.     

Investigation of lipid bodies in a colon carcinoma cell line by confocal Raman microscopy

Objective

The investigation of living cells under physiological conditions requires sensitive, sophisticated and in particular, non-invasive methods. Raman spectroscopy provides vibrational information about the sample. Combined with high-resolution confocal microscopy, it allows a complete Raman spectrum to be recorded at every confocal image point. This technique was applied here for the investigation of lipid bodies in a colon carcinoma cell line.

Materials and methods

The colorectal adenocarcinoma cell line Caco-2 and the rat intestine epithelial cell line IEC-6 were analysed with the confocal Raman microscope alpha300 R (WITec GmbH, Germany), using a frequency-doubled Nd:YAG laser at 532 nm and 10 mW for excitation. The use of a water immersion lens (63×, NA 1.0) allowed a lateral resolution in the sub-micrometer range. Raman images of cells were generated from the data sets by integrating over specific Raman bands. Mapping of the C–H stretching band (2800–3030 cm⁻¹) allowed for the visualisation of the whole cell, whereas the automated statistical evaluation of all spectra by k-means cluster analysis resulted in spectral unmixed images which provided an insight into the chemical composition of the sample.

Results

With the described method, it was possible to visualise the distribution of different cellular biomarkers. Lipid bodies, in particular, which are reported to be present in increased numbers in colorectal cancer cells as compared to normal tissue, could be characterised and localised. A quantitative approach was developed to assess the fraction of lipid bodies in the total cell area. This method was applied for comparison between malignant and non-malignant cell lines. The fraction of lipid bodies turned out to be significantly higher in malignant (13.8%, n = 21) than in non-malignant cells (1.8%, n = 16).

Conclusion

Confocal Raman microscopy is shown to be a powerful method for the investigation of lipid bodies.     

Non-contact discrimination of human bone marrow-derived mesenchymal stem cells and fibroblasts using Raman spectroscopy

Background

Human bone marrow-derived mesenchymal stem cells (BM-MSCs) are a promising cell source for regenerative medical applications because they can be easily isolated and expanded ex vivo. However, due to the lack of specific cell surface markers, it is difficult to identify whether ex vivo isolated BM-MSC populations are free of stromal fibroblasts. Bright-field microscopical analyses are insufficient to determine fibroblastic contaminations since these two cell types have similar cell morphologies.

Materials and methods

We employed and compared traditional flow cytometric (FACS) analysis, in vitro differentiation assays, and Raman spectroscopy to distinguish between human BM-MSCs and fibroblasts.

Results

We found that FACS analysis, utilizing previously described fibroblast-identifying antibodies, was inadequate in separating stromal fibroblasts from BM-MSCs as over 75% of the BM-MSCs shared these antigens. In vitro differentiation assays revealed that, in contrast to fibroblasts, BM-MSCs could be successfully differentiated into adipocytes, osteoblasts and chondrocytes. Using this method it was possible to discriminate between the two cell types. However, the need for prolonged in vitro culture periods of up to 4 weeks is a major disadvantage of this test method. Raman spectroscopy, a non-contact technique measuring the wavelength and intensity of inelastic scattered light from molecules by employing high-power near-infrared lasers, distinguished ultra-fast between BM-MSCs and fibroblasts (integration time of 100 s/cell).

Conclusion

Based on the results, we conclude that Raman spectroscopy is a suitable tool for the rapid detection of fibroblastic contaminations in BM-MSC cultures.