In the present study, an innovative and highly efficient near-infrared hyperspectral imaging (NIR-HSI) method is proposed to provide spectral maps able to reveal collagen distribution in large-size ...bones, also offering semi-quantitative estimations. A recently introduced method for the construction of chemical maps, based on Normalized Difference Images (NDI), is declined in an innovative approach, through the exploitation of the NDI values computed for each pixel of the hyperspectral image to localize collagen and to extract information on its content by a direct comparison with known reference samples. The developed approach addresses an urgent issue of the analytical chemistry applied to bioarcheology researches, which rely on well-preserved collagen in bones to obtain key information on chronology, paleoecology and taxonomy. Indeed, the high demand for large-sample datasets and the consequent application of a wide variety of destructive analytical methods led to the considerable destruction of precious bone samples. NIR-HSI pre-screening allows researchers to properly select the sampling points for subsequent specific analyses, to minimize costs and time and to preserve integrity of archaeological bones (which are available in a very limited amount), providing further opportunities to understand our past.
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•Near-infrared hyperspectral imaging for localising collagen in ancient bones.•Computation of normalized difference images for high-throughput chemical mapping.•Relative estimation of collagen content within a sample and between samples.•Efficient non-destructive strategy to select sampling areas in entire specimens.•Green and rapid screening method which saves bones for further analyses.
The identification of organic dyes is a challenging task in all the fields such as the forensic and conservation sciences, especially in cases where the amount of sample is extremely small. In this ...paper we propose a new enhanced FTIR method (MU-ATR metal underlayer ATR spectroscopy), which we believe is the first of its kind, for the analysis of a few ng of dyes. With this method, dyed fiber micro-extracts can be analyzed using a commercial FTIR microscope with a fixed incident angle, obtaining the same separation between the different classes of dyes investigated as we obtained analyzing pure dyes in transmission mode.
Moreover, the new enhancement method has been validated on a real sample dated back to the 1893, showing how it can be promising for the analysis of trace amounts of organic substances in artistic samples such as dyes in paintings or textiles, varnishes and organic residues on archaeological objects.
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•MU-ATR mode is a new FTIR enhanced method.•Simulation and experimental data were acquired to understand the enhancement effect.•MU-ATR spectra can be acquired on few ng of dyes.•Classes of dyes were distinguished analyzing 10−5 M solutions in MU-ATR.•MU-ATR has been validated on a real sample.
The benefits for elderly individuals of regular participation in both cardiovascular and resistance-training programmes are great. Health benefits include a significant reduction in risk of coronary ...heart disease, diabetes mellitus and insulin resistance, hypertension and obesity as well as improvements in bone density, muscle mass, arterial compliance and energy metabolism. Additionally, increases in cardiovascular fitness (maximal oxygen consumption and endurance), muscle strength and overall functional capacity are forthcoming allowing elderly individuals to maintain their independence, increase levels of spontaneous physical activity and freely participate in activities associated with daily living. Taken together, these benefits associated with involvement in regular exercise can significantly improve the quality of life in elderly populations. It is noteworthy that the quality and quantity of exercise necessary to elicit important health benefits will differ from that needed to produce significant gains in fitness. This review describes the current recommendations for exercise prescriptions for the elderly for both cardiovascular and strength/resistance-training programmes. However, it must be noted that the benefits described are of little value if elderly individuals do not become involved in regular exercise regimens. Consequently, the major challenges facing healthcare professionals today concern: (i) the implementation of educational programmes designed to inform elderly individuals of the health and functional benefits associated with regular physical activity as well as how safe and effective such programmes can be; and (ii) design interventions that will both increase involvement in regular exercise as well as improve adherence and compliance to such programmes.
This study shows that state-of-the-art liquid chromatography (LC) and mass spectrometry (MS) can be used for rapid verification of identity and characterization of sequence variants and ...posttranslational modifications (PTMs) for antibody products. A candidate biosimilar IgG1 monoclonal antibody (mAb) was compared in detail to a commercially available innovator product. Intact protein mass, primary sequence, PTMs, and the micro-differences between the two mAbs were identified and quantified simultaneously. Although very similar in terms of sequences and modifications, a mass difference observed by LC-MS intact mass measurements indicated that they were not identical. Peptide mapping, performed with data independent acquisition LC-MS using an alternating low and elevated collision energy scan mode (LC-MS
E
), located the mass difference between the biosimilar and the innovator to a two amino acid residue variance in the heavy chain sequences. The peptide mapping technique was also used to comprehensively catalogue and compare the differences in PTMs of the biosimilar and innovator mAbs. Comprehensive glycosylation profiling confirmed that the proportion of individual glycans was different between the biosimilar and the innovator, although the number and identity of glycans were the same. These results demonstrate that the combination of accurate intact mass measurement, released glycan profiling, and LC-MS
E
peptide mapping provides a set of routine tools that can be used to comprehensively compare a candidate biosimilar and an innovator mAb.
We report the development of innovative embedding procedures for the analysis of paint cross sections by attenuated total reflection (ATR) Fourier transform IR microscopy. This technique was chosen ...because it is widely employed for the characterization and spatial location of organic and inorganic components in artistic samples. Moreover, the performance of the technique may be critically affected by sample preparation in terms of surface morphology and the presence of contamination. First, we evaluated the use of KBr as a barrier to contamination by the embedding synthetic medium. In this way, the sample cross section can be polished by means of a sample holder, which allows a controlled pressure to be applied to the sample, thus improving the reproducibility and quality of the surface cross section. In addition, argon ion milling was used for the polishing of samples embedded in KBr, and provided very promising results in terms of surface planarity and reduction of superficial contamination by KBr. Finally, the use of NaCl as an alternative to KBr was proposed thanks to its advantages in terms of hygroscopicity, cost, and toxicity. In addition, cross sections embedded in NaCl were characterized by greater hardness, a feature that allowed us to obtain improved contact with the ATR crystal.
Monoclonal antibodies are typically glycosylated at asparagine residues in the Fc domain, and glycosylation heterogeneity at the Fc sites is well known. This paper presents a method for rapid ...analysis of glycosylation profile of the therapeutic monoclonal antibody trastuzumab from different production batches using electrospray quadrupole ion-mobility time-of-flight mass spectrometry (ESI-Q-IM-TOF). The global glycosylation profile for each production batch was obtained by a fast LC-MS analysis, and comparisons of the glycoprofiles of trastuzumab from different lots were made based on the deconvoluted intact mass spectra. Furthermore, the heterogeneity at each glycosylation site was characterized at the reduced antibody level and at the isolated glycopeptide level. The glycosylation site and glycan structures were confirmed by performing a time-aligned-parallel fragmentation approach using the unique dual-collision cell design of the instrument and the incorporated ion-mobility separation function. Four different production batches of trastuzumab were analyzed and compared in terms of global glycosylation profiles as well as the heterogeneity at each glycosylation site. The results show that each batch of trastuzumab shares the same types of glycoforms but relative abundance of each glycoforms is varied.
Glycosylation heterogeneity of different production batches of a therapeutic monoclonal antibody was investigated using quadrupole-ion-mobility mass spectrometry.
The characterization of the organic components in a complex, multilayered paint structure is fundamental for studying painting techniques and for authentication and restoration purposes. ...Proteinaceous materials, such as animal glue, are of particular importance since they are widely used as binders, adhesives and for gilding. Even though proteins are usually detected by chromatographic and proteomic techniques, immunological methods represent an alternative powerful approach to protein analysis thanks to the high specificity of antigen–antibody reactions. Our previous studies demonstrated that ovalbumin and casein could be localized in paint cross-sections with high sensitivity and good spatial resolution (i.e. within the single painting layers) by using chemiluminescent (CL) immunochemical microscope imaging. In the present research work, we describe for the first time the immunolocalization of collagen (the main protein of animal glue) in paint cross-sections by CL imaging microscopy. Two different analytical protocols have been developed, allowing either the detection of collagen or the simultaneous detection of collagen and ovalbumin in the same paint sample. The assays were used to detect collagen and ovalbumin in cross-sections from model samples and historical paintings (a wall painting dated to 1773–1774 and a painted wood panel of the Renaissance period) in order to achieve information on paint techniques and past restoration interventions.
Figure
Left
Reflected light image of a cross-section of a sample taken from a Renaissance painted wood panel.
Right
Localization of the proteins collagen (from animal glue) and ovalbumin in a painting cross-section assessed by multiplexed chemiluminescence immunochemical imaging (the chemiluminescent signals corresponding to collagen and ovalbumin are displayed in shades of
blue
and
red
, respectively)
To study degradation processes occurring on painting materials, the use of high-resolution micro-analytical techniques is highly requested since it provides a detailed identification and localisation ...of both the original and deteriorated ingredients. Among the various pigments recently studied, the characterisation of verdigris has received a major interest. This pigment has not a unique chemical formula, but its composition depends on the recipe employed for its manufacturing. Moreover, verdigris paints are not stable and are subject to a colour change from blue-green to green, which occurs in the first few months after the application. In this paper, we focused our attention on the use of ATR-FTIR mapping as a useful method to identify verdigris secondary products and pathways. Several mock-ups and real samples have been analysed, and the correlation among the detected compounds and their spatial location, obtained by the application of ATR-FTIR microscopy in mapping mode, allowed formulating some hypotheses on the degradation pattern of verdigris, which may feed the discussion on the transformation and stability of this pigment. From an analytical point of view, we showed how FTIR mapping approaches may be extremely useful both for the identification of compounds in complex matrix in which single spectra may limit the exhaustive characterisations due to bands overlapping and for the study of degradation pathways by taking into consideration the relative distribution of degradation products.
•Laser pyrolysis allows the localisation of organic substances in paint cross sections.•Natural resins may be distinguished and localised.•Synthetic colourants may be distinguished and localised.
The ...ability to analyse layers within a paint sample is important both for historical studies as well as determining the most suitable method for restoration. The difficulty in physically separating these very thin layers makes detailed analysis difficult, and a combination of destructive and non-destructive techniques are needed. Laser pyrolysis/gas chromatography - mass spectrometry (LaPy/GC–MS) is a technique that has proved successful in the analysis of geological samples which pose the same difficulties for analysis, but it has not yet been tried on paint cross sections. This study presents preliminary results of applying LaPy/GC–MS on standard mock-ups for identification of organic compounds used as varnishes, binders and colourants.
The technique was shown to be particularly effective for the identification and localisation of natural resins and synthetic colourants. Moreover, the integration between this technique and other stratigraphical analyses may allow the complete characterisation of the composition of organic substances in paint cross-sections.
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