Protein lipoxidation:detection strategies and challenges

Enzymatic and non-enzymatic lipid metabolism can give rise to reactive species that may covalently modify cellular or plasma proteins through a process known as lipoxidation. Under basal conditions, protein lipoxidation can contribute to normal cell homeostasis and participate in signaling or adaptive mechanisms, as exemplified by lipoxidation of Ras proteins or of the cytoskeletal protein vimentin, both of which behave as sensors of electrophilic species. Nevertheless, increased lipoxidation under pathological conditions may lead to deleterious effects on protein structure or aggregation. This can result in impaired degradation and accumulation of abnormally folded proteins contributing to pathophysiology, as may occur in neurodegenerative diseases. Identification of the protein targets of lipoxidation and its functional consequences under pathophysiological situations can unveil the modification patterns associated with the various outcomes, as well as preventive strategies or potential therapeutic targets. Given the wide structural variability of lipid moieties involved in lipoxidation, highly sensitive and specific methods for its detection are required. Derivatization of reactive carbonyl species is instrumental in the detection of adducts retaining carbonyl groups. In addition, use of tagged derivatives of electrophilic lipids enables enrichment of lipoxidized proteins or peptides. Ultimate confirmation of lipoxidation requires high resolution mass spectrometry approaches to unequivocally identify the adduct and the targeted residue. Moreover, rigorous validation of the targets identified and assessment of the functional consequences of these modifications are essential. Here we present an update on methods to approach the complex field of lipoxidation along with validation strategies and functional assays illustrated with well-studied lipoxidation targets.

Simultaneous measurement for liquid level and refractive index based on all-fiber modal interferometer

A simple fiber sensor capable of simultaneous measurement of liquid level and refractive index (RI) is proposed and experimentally demonstrated. The sensing head is an all-fiber modal interferometer manufactured by splicing an uncoated single-mode fiber with two short sections of multimode fiber. The interference pattern experiences blue shift along with an increase of axial strain and surrounding RI. Owing to the participation of multiple cladding modes with different sensitivities, the height and RI of the liquid could be simultaneously measured by monitoring two dips of the transmission spectrum. Experimental results show that the liquid level and RI sensitivities of the two dips are 245.7 pm/mm, -38 nm/RI unit (RIU), and 223.7 pm/mm, -62 nm/RIU, respectively. The approach has distinctive advantages of easy fabrication, low cost, and high sensitivity for liquid level detection with the capability of distinguishing the RI variation simultaneously. © 2013 Copyright Taylor and Francis Group, LLC.

Refractive index and temperature sensitivity characteristics of a micro-slot fiber Bragg grating

Fabrication and characterization of a UVinscribed fiber Bragg grating (FBG) with a micro-slot liquid core is presented. Femtosecond (fs) laser patterning/chemical etching technique was employed to engrave a micro-slot with dimensions of 5.74 μm(h) × 125 μm(w) × 1388.72 μm(l) across the whole grating. The device has been evaluated for refractive index (RI) and temperature sensitivities and exhibited distinctive thermal response and RI sensitivity beyond the detection limit of reported fiber gratings. This structure has not just been RI sensitive, but also maintained the robustness comparing with the bare core FBGs and long-period gratings with the partial cladding etched off. © 2012 Optical Society of America.

Detection and quantification of protein oxidation in sarcopenic models:a mass spectrometry study

Oxidised biomolecules in aged tissue could potentially be used as biomarkers for age-related diseases; however, it is still unclear whether they causatively contribute to ageing or are consequences of the ageing process. To assess the potential of using protein oxidation as markers of ageing, mass spectrometry (MS) was employed for the identification and quantification of oxidative modifications in obese (ob/ob) mice. Lean muscle mass and strength is reduced in obesity, representing a sarcopenic model in which the levels of oxidation can be evaluated for different muscular systems including calcium homeostasis, metabolism and contractility. Several oxidised residues were identified by tandem MS (MS/MS) in both muscle homogenate and isolated sarcoplasmic reticulum (SR), an organelle that regulates intracellular calcium levels in muscle. These modifications include oxidation of methionine, cysteine, tyrosine, and tryptophan in several proteins such as sarcoplasmic reticulum calcium ATPase (SERCA), glycogen phosphorylase, and myosin. Once modifications had been identified, multiple reaction monitoring MS (MRM) was used to quantify the percentage modification of oxidised residues within the samples. Preliminary data suggests proteins in ob/ob mice are more oxidised than the controls. For example SERCA, which constitutes 60-70% of the SR, had approximately a 2-fold increase in cysteine trioxidation of Cys561 in the obese model when compared to the control. Other obese muscle proteins have also shown a similar increase in oxidation for various residues. Further analysis with complex protein mixtures will determine the potential diagnostic use of MRM experiments for analysing protein oxidation in small biological samples such as muscle needle biopsies.

Localized surface plasmon fiber device coated with carbon nanotubes for the specific detection of CO2

We explored the potential of a carbon nanotube (CNT) coating working in conjunction with a recently developed localized surface plasmon (LSP) device (based upon a nanostructured thin film consisting of of nano-wires of platinum) with ultra-high sensitivity to changes in the surrounding index. The uncoated LSP sensor’s transmission resonances exhibited a refractive index sensitivity of Δλ/Δn ~ -6200nm/RIU and ΔΙ/Δn ~5900dB/RIU, which is the highest reported spectral sensitivity of a fiber optic sensor to bulk index changes within the gas regime. The complete device provides the first demonstration of the chemically specific gas sensing capabilities of CNTs utilizing their optical characteristics. This is proven by investigating the spectral response of the sensor before and after the adhesion of CNTs to alkane gases along with carbon dioxide. The device shows a distinctive spectral response in the presence of gaseous CO2 over and above what is expected from general changes in the bulk refractive index. This fiber device yielded a limit of detection of 150ppm for CO2 at a pressure of one atmosphere. Additionally the adhered CNTs actually reduce sensitivity of the device to changes in bulk refractive index of the surrounding medium. The polarization properties of the LSP sensor resonances are also investigated and it is shown that there is a reduction in the overall azimuthal polarization after the CNTs are applied. These optical devices offer a way of exploiting optically the chemical selectivity of carbon nanotubes, thus providing the potential for real-world applications in gas sensing in many inflammable and explosive environments. © (2015) COPYRIGHT Society of Photo-Optical Instrumentation Engineers (SPIE). Downloading of the abstract is permitted for personal use only.

Optimisation of chromatographic separation of oxidised phospholipids and detection with targeted approaches provides greater coverage of the oxidised lipidome

Phospholipid oxidation by adventitious damage generates a wide variety of products with potentially novel biological activities that can modulate inflammatory processes associated with various diseases. To understand the biological importance of oxidised phospholipids (OxPL) and their potential role as disease biomarkers requires precise information about the abundance of these compounds in cells and tissues. There are many chemiluminescence and spectrophotometric assays available for detecting oxidised phospholipids, but they all have some limitations. Mass spectrometry coupled with liquid chromatography is a powerful and sensitive approach that can provide detailed information about the oxidative lipidome, but challenges still remain. The aim of this work is to develop improved methods for detection of OxPLs by optimisation of chromatographic separation through testing several reverse phase columns and solvent systems, and using targeted mass spectrometry approaches. Initial experiments were carried out using oxidation products generated in vitro to optimise the chromatography separation parameters and mass spectrometry parameters. We have evaluated the chromatographic separation of oxidised phosphatidylcholines (OxPCs) and oxidised phosphatidylethanolamines (OXPEs) using C8, C18 and C30 reverse phase, polystyrene – divinylbenzene based monolithic and mixed – mode hydrophilic interaction (HILIC) columns, interfaced with mass spectrometry. Our results suggest that the monolithic column was best able to separate short chain OxPCs and OxPEs from long chain oxidised and native PCs and PEs. However, variation in charge of polar head groups and extreme diversity of oxidised species make analysis of several classes of OxPLs within one analytical run impractical. We evaluated and optimised the chromatographic separation of OxPLs by serially coupling two columns: HILIC and monolith column that provided us the larger coverage of OxPL species in a single analytical run.

Highly sensitive refractive index sensor based on cascaded microfiber knots with Vernier effect

We propose and experimentally demonstrate a refractive index (RI) sensor based on cascaded microfiber knot resonators (CMKRs) with Vernier effect. Deriving from high proportional evanescent field of microfiber and spectrum magnification function of Vernier effect, the RI sensor shows high sensitivity as well as high detection resolution. By using the method named "Drawing-Knotting-Assembling (DKA)", a compact CMKRs is fabricated for experimental demonstration. With the assistance of Lorentz fitting algorithm on the transmission spectrum, sensitivity of 6523nm/RIU and detection resolution up to 1.533 x 10-7 RIU are obtained in the experiment which show good agreement with the numerical simulation. The proposed all-fiber RI sensor with high sensitivity, compact size and low cost can be widely used for chemical and biological detection, as well as the electronic/magnetic field measurement

High-sensitivity refractive index sensor based on large-angle tilted fiber grating with carbon nanotube deposition

This paper presents a highly sensitive ambient refractive index (RI) sensor based on 81° tilted fiber grating (81°-TFG) structure UV-inscribed in standard telecom fiber (62.5μm cladding radius) with carbon nanotube (CNT) overlay deposition. The sensing mechanism is based on the ability of CNT to induce change in transmitted optical power and the high sensitivity of 81°-TFG to ambient refractive index. The thin CNT film with high refractive index enhances the cladding modes of the TFG, resulting in the significant interaction between the propagating light and the surrounding medium. Consequently, the surrounding RI change will induce not only the resonant wavelength shift but also the power intensity change of the attenuation band in the transmission spectrum. Result shows that the change in transmitted optical power produces a corresponding linear reduction in intensity with increment in RI values. The sample shows high sensitivities of ∼207.38nm/RIU, ∼241.79nm/RIU at RI range 1.344-1.374 and ∼113.09nm/RIU, ∼144.40nm/RIU at RI range 1.374-1.392 (for X-pol and Y-pol respectively). It also shows power intensity sensitivity of ∼ 65.728dBm/RIU and ∼ 45.898 (for X-pol and Y-pol respectively). The low thermal sensitivity property of the 81°-TFG offers reduction in thermal cross-sensitivity and enhances specificity of the sensor.