Assessment by Matrix‐Assisted Laser Desorption/Ionization Time‐of‐Flight Mass Spectrometry of the Effects of Preanalytical Variables on Serum Peptidome Profiles Following Long‐Term Sample Storage

Purpose

Human serum and plasma are often used as clinical specimens in proteomics analyses, and peptidome profiling of human serum is a promising tool for identifying novel disease‐associated biomarkers. Matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry (MALDI‐TOF MS) is widely used for peptidomic biomarker discovery. Careful sample collection and handling are required as either can have a profound impact on serum peptidome patterns, yet the effects of preanalytical variables on serum peptidome profiles have not been completely elucidated. The present study investigated the effects of preanalytical variables, including storage temperature, duration (up to 12 months), and thawing methods, on MALDI‐TOF MS‐based serum peptidome patterns.

Experimental design

Aliquots of serum samples were pretreated with weak cation exchanger magnetic beads using an automated ClinProtRobot system and then analyzed by MALDI‐TOF MS.

Results

A number of significant differences in peak intensities were observed depending on sample processing variables.

Conclusions and clinical relevance

These peaks can be used as sample quality markers to assess the effects of long‐term storage on serum peptidome profiles using MALDI‐TOF MS.

     

Two dimensional gel electrophoresis of apolipoprotein C‐III and MALDI‐TOF MS are complementary techniques for the study of combined defects in N‐ and mucin type O‐glycan biosynthesis

In the field of diseases related to glycosylation disorders, congenital defects associated with abnormalities in both O‐ and N‐glycosylation of proteins constitute arising novel entities. Defects in subunits of the conserved oligomeric Golgi protein complex have been shown to be involved in an important part of previously unsolved CDG type II combining abnormalities in both mucin type core1 O‐ and N‐glycans; furthermore, recent studies revealed that autosomal recessive cutis laxa type II could also be associated with such combined glycosylation defects. Based on the studies of serum samples from three patients including a case of cutis laxa, we present here evidence that 2‐DE of apolipoprotein C‐III in combination with MALDI‐TOF‐MS analysis of serum O‐ and N‐glycans allow the detection and the biochemical characterization of these newly recognized glycosylation disorders.     

Translating N‐Glycan Analytical Applications into Clinical Strategies for Ovarian Cancer

Protein glycosylation, particularly N‐linked glycosylation, is a complex posttranslational modification (PTM), which plays an important role in protein folding and conformation, regulating protein stability and activity, cell–cell interaction, and cell signaling pathways. This review focuses on analytical techniques, primarily MS‐based techniques, to qualitatively and quantitatively assess N‐glycosylation while successfully characterizing compositional, structural, and linkage features with high specificity and sensitivity. The analytical techniques explored in this review include LC–ESI–MS/MS and MALDI time‐of‐flight MS (MALDI‐TOF‐MS), which have been used to analyze clinical samples, such as serum, plasma, ascites, and tissue. Targeting the aberrant N‐glycosylation patterns observed in MALDI–MS imaging (MSI) offers a platform to visualize N‐glycans in tissue‐specific regions. The studies on the intra‐patient (i.e., a comparison of tissue‐specific regions from the same patient) and inter‐patient (i.e., a comparison of tissue‐specific regions between different patients) variation of early‐ and late‐stage ovarian cancer (OC) patients identify specific N‐glycan differences that improve understanding of the tumor microenvironment and potentially improve therapeutic strategies for the clinic.     

Identification of melanoma‐specific serological markers using proteomic analyses

The discovery of novel melanoma markers for not only early detection but also monitoring disease status is promising to improve the clinical outcome of patients. In the present study, we performed proteomic comparative analysis of plasma proteins between healthy volunteers and melanoma patients using NanoLC and MALDI‐TOF‐MS. As a result, we were successful in identifying nine proteins that were specifically expressed in melanoma plasma compared with healthy plasma, most of which had not previously been identified as plasma markers of melanoma. The mRNA expression levels of four proteins [pro‐platelet basic protein precursor (PPBP), serum amyloid A2 (SAA2), complement factor H‐related protein 1 precursor (FHR1), inter‐alpha‐trypsin inhibitor heavy chain H4 precursor (IAIH4)] were prominently up‐regulated in several melanoma cell lines compared with melanocytes. Moreover, two proteins (PPBP, SAA) were shown to be expressed in tumor specimens from melanoma patients. In the survival time analysis regarding melanoma patients, the semi‐quantified plasma PPBP levels were statistically negatively correlated with the survival time. Most interestingly, the significant survival benefit was seen in low PBPP level group (< index 20) versus high level (≥ index 20) group. The results suggest that PPBP might be a novel promising serological marker and a prognostic factor specific to melanomas.     

Automated serum peptide profiling using novel magnetic C18 beads off-line coupled to MALDI-TOF-MS

Serum peptide profiling by MS is an emerging approach for disease diagnosis and biomarker discovery. A magnetic bead‐based method for off‐line serum peptide capture coupled to MALDI‐TOF‐MS has been recently introduced. However, the reagents are not available to the general scientific community. Here, we developed a protocol for serum peptide capture using novel magnetic C18 beads, and automated the procedure on a high‐throughput magnetic particle processor. We investigated bead equilibration, peptide binding and peptide elution conditions. The method is evaluated in terms of peaks counts and reproducibility of ion intensities in control serum. Overall, the DynaBead‐RPC18‐based serum sample processing protocol reported here is reproducible, robust and allows for the detection of ?200 peptides at m/z 800–4000 of serum that was allowed to clot for 1 h. The average intra‐experiment %CV of normalized ion intensities for crude serum and 0.5% TFA/0.15% n‐octyl glucoside‐treated serum, respectively, were 12% (range 2–38%) and 10% (3–21%) and the inter‐experiment %CVs were 24% (10–53%) and 31% (10–59%). Importantly, this method can be used for serum peptide profiling by anyone in possession of a MALDI‐TOF instrument. In conjunction with the KingFisher® 96, the whole serum peptide capture procedure is high‐throughput (?20 min per isolation of 96 samples in parallel), thereby facilitating large‐scale disease profiling studies.     

Towards stable diagnostic setups in clinical proteomics: Absolute quantitation of peptide biomarkers using MALDI‐TOF‐MS

In routine clinical diagnostics, peptide biomarkers are most commonly quantified using immunological techniques but these methods often lack sensitivity and/or specificity. Hence, quantitative mass spectrometry detection is desirable as an alternative diagnostic tool. To date, quantitative mass spectrometry is mostly based on ESI‐MS coupled to LC, requiring highly sophisticated instrumentation and knowledge and is time consuming and expensive. In contrast, MALDI‐TOF‐MS is a very simple, sensitive and rapid method for the detection of peptide biomarkers. However, the infeasibility of absolute quantification has been a tremendous handicap to the use of MS in stable clinical diagnostics. Here, we describe the development of a technical platform based on ClinProt particles and heavy‐isotope internal peptide standards for the fast and reliable preparation of samples. This combines the advantages of MALDI‐TOF as a read‐out system with absolute quantitation of peptide biomarkers. As a proof‐of‐concept, this platform was successfully employed for the absolute determination of the concentration of the highly abundant serum peptide des‐Ala‐Fibrinopeptide A in 45 serum samples from healthy donors. Such technology essentially contributes to the development of a stable MALDI‐TOF‐MS‐based clinical assay.     

Changes of serum‐associated fucosylated glycoproteins and changes in glycosylation of IgA in human cirrhosis

Many modifications in N‐glycosylation have been demonstrated in hepatic cirrhosis. These modifications correspond to an increase of a bisecting core alpha(1,6)‐fucosylated biantennary glycan, an increase in core fucosylation, and the presence of an important population of neutral oligosaccharides in human serum of cirrhotic patients. In this study, a glycoproteomic approach which consists of lectin affinity chromatography, MALDI‐TOF MS for the characterization of N‐glycans released from glycoproteins, one‐ and 2‐D PAGE, electrospray ionization quadrupole ion trap (ESI‐QIT) MS was used to identify serum fucosylated glycoproteins related to cirrhosis. Employing this method, we have shown that IgA is one of the major proteins that is responsible of the glycosylation modifications observed in the serum N‐glycome of cirrhotic patients. To our knowledge, this is the first time that aberrant N‐glycosylation of IgA in cirrhosis is described.     

The multiple loop antenna for enhanced readability performances of near‐field UHF RFID applications

This research is concerned with a multiple loop antenna applicable to near field ultra‐high frequency (UHF) radio frequency identification (RFID). The proposed multi‐loop antenna is configured to induce the robust and even magnetic field distributions in the H_x, H_y, and H_z orientations so as to achieve the enhanced readability performances in all directions (i.e., x, y, and z). Simulations were carried out using CST Microwave Studio to determine the impedance bandwidth (|S₁₁|H_x‐, H_y‐, and H_z‐oriented magnetic field distributions. A prototype antenna of 14 cm × 16 cm × 0.6 mm (W × L × H) in overall dimension was subsequently fabricated on an FR4 substrate connected to a coaxial cable. In this research, the H_x‐, H_y‐, and H_z‐oriented magnetic field distributions of the prototype antenna were measured in the x‐axis, y‐axis, and x–y plane and are in good agreement with the simulation results. The measured readability performances in the x‐, y‐, and z‐directions in which seven near field UHF RFID tags were deployed unobstructed (i.e., in open air) are respectively 25.27%, 31.73%, and 85.43%. Furthermore, the performances on the antenna readability with the tags attached to the microcentrifuge tubes are 30.55%, 25.90%, and 69.09% for the x‐, y‐, and z‐directions. © 2016 Wiley Periodicals, Inc. Int J RF and Microwave CAE 26:402–417, 2016.     

Mass Spectrometric Analysis of Cerebrospinal Fluid Ubiquitin in Alzheimer's Disease and Parkinsonian Disorders

1 Purpose

Dysfunctional proteostasis, with decreased protein degradation and an accumulation of ubiquitin into aggregated protein inclusions, is a feature of neurodegenerative diseases. Identifying new potential biomarkers in cerebrospinal fluid (CSF) reflecting this process could contribute important information on pathophysiology.

2 Experimental design

A developed method combining SPE and PRM‐MS is employed to monitor the concentration of ubiquitin in CSF from subjects with Alzheimer's disease (AD), Parkinson's disease (PD), and progressive supranuclear palsy (PSP). Four independent cross‐sectional studies are conducted, studies 1–4, including controls (n = 86) and participants with AD (n = 60), PD (n = 15), and PSP (n = 11).

3 Results

The method shows a repeatability and intermediate precision not exceeding 6.1 and 7.9%, respectively. The determined LOD is 0.1 nm and the LOQ range between 0.625 and 80 nm . The CSF ubiquitin concentration is 1.2–1.5‐fold higher in AD patients compared with controls in the three independent AD‐control studies (Study 1, p < 0.001; Study 2, p < 0.001; and Study 3, p = 0.003). In the fourth study, there is no difference in PD or PSP, compared to controls.

4 Conclusion and clinical relevance

CSF ubiquitin may reflect dysfunctional proteostasis in AD. The described method can be used for further exploration of ubiquitin as a potential biomarker in neurodegenerative diseases.     

Measurement of whole‐body vibration exposure from unsuspended cabin tractor semi‐trailers

Indian tractor semi‐trailer drivers are exposed to mechanical whole‐body vibration during their work. Some drivers suffer from low‐back pain from this vibration. However, there is no evidence of a relationship between the whole‐body vibration from tractor semi‐trailers and low‐back pain or occupational disease because of the lack of investigations. A field study was conducted to characterize the health risks associated with driving tractor semi‐trailers. Studies were conducted at different loadings and on different road surfaces as well as at different speeds, with the vibrations measured at the driver–seat interface on x‐longitudinal, y‐transverse and z‐vertical axes. The vibrations were compared with the health‐risk guidance according to Annex B of ISO 2631‐1 (ISO 2631/1, 1997). The findings of this study indicated that Indian tractor semi‐trailer drivers should not operate continuously more than 4 h a day under current working conditions. © 2011 Wiley Periodicals, Inc.     

2‐D ALGEBRAIC TEST FOR ROBUST STABILITY OF QUASIPOLYNOMIALS WITH INTERVAL PARAMETERS

Determining the robust stability of interval quasipolynomials leads to a NP problem: an enormous number of testing edge polynomials. This paper develops an efficient approach to reducing the number of testing edge polynomials. This paper solves the stability test problem of interval quasipolynomials by transforming interval quasipolynomials into two‐dimensional (2‐D) interval polynomials. It is shown that the robust stability of an interval 2‐D polynomial can ensure the stability of the quasipolynomial, and the algebraic test algorithm for 2‐D s‐z interval polynomials is provided. The stability of 2‐D s‐z vertex polynomials and 2‐D s‐z edge polynomials were tested by using a Schur Table of complex polynomials.     

Poststress RF‐drifts of dual‐band LC VCO in 0.18‐μm CMOS technology

This article studies the RF‐property of a dual‐band voltage‐controlled oscillator (VCO). The designed circuit consists of a dual‐resonance LC resonator and a Colpitts negative resistance cell. The dual‐resonance LC resonator comprises a series‐tuned LC resonator and a parallel resonant resonator. The proposed VCO has been implemented with the TSMC 0.18 μm 1P6M CMOS technology. The VCO can generate differential signals in the frequency range of 3.0–3.37 GHz and 6.95–7.40 GHz with core power consumption of 10.08 and 10.24 mW at the dc drain‐source bias V_DD of 1.4 V, respectively. The die area of the dual‐band VCO is 0.485 × 0.800 mm². The circuit was operated at V_DD = 3 V for 8 h and significant drift in RF parameters was found. © 2013 Wiley Periodicals, Inc. Int J RF and Microwave CAE 24:243–248, 2014.     

White LEDs using the sharp β‐sialon: Eu phosphor and Mn‐doped red phosphor for wide‐color gamut display applications

The sharp β‐sialon (Si_6‐zAl_zO_zN_8‐z : 0 < z < 0.1):Eu green phosphor, combining with a blue LED and CaAlSiN₃:Eu red phosphor, is suitable for the wide‐color gamut white LEDs backlighting system, because of its sharp and asymmetric emission spectrum shape. However, the color gamut and the brightness of the aforementioned display is restricted because of the wide emission band of the CaAlSiN₃:Eu red phosphor. In this work, we used K₂SiF₆:Mn as an alternate red phosphor, which has a sharp emission spectrum. The display with the white LED using sharp β‐sialon:Eu and K₂SiF₆:Mn shows a wide‐color gamut, which covers the hole NTSC triangle. The use of K₂SiF₆:Mn enables to realize not only a wider color gamut but also a higher brightness of displays, compared with the use of CaAlSiN₃:Eu. Furthermore, it is confirmed that the white LED using sharp β‐sialon:Eu and K₂SiF₆:Mn is stable against temperature and also durable under the accelerated drive conditions.     

A 4‐bit ultra‐wideband complementary metal‐oxide‐semiconductor attenuator with low root‐mean‐square amplitude error

This article presents the 4‐bit ultra‐wideband complementary metal‐oxide‐semiconductor (CMOS) attenuator in a standard 0.18‐μm CMOS process. This design adopts switched bridge‐T type topologies for each attenuation bit. Based on insertion losses and input P_1‐dB considerations, the circuit performances can be optimized by the proper bit ordering arrangement. Therefore, the bit ordering 0.5‐4‐2‐1 dB is employed in the 4‐bit attenuator. Moreover, series inductors are added between each bit to further improve the input and output return losses. Measured results demonstrate that the attenuation range of the circuit is 7.5 dB with 0.5 dB step and the root‐mean‐square (RMS) amplitude error is between 0.11 and 0.13 dB from 3.1 to 10.8 GHz. The differences between simulated and measured RMS amplitude errors are less than 0.2 dB, which demonstrates the good agreement and feasibility of the design concept. The measured input P_1‐dB is 15 dBm at 5 GHz and the chip area is 1.12 mm² including all testing pads.     

Finite‐time boundedness of switched systems with time‐varying delays via sampled‐data control

In the framework of sampled‐data control, finite‐time boundedness (FTB) of switched systems with time‐varying delays is investigated. Sufficient conditions for FTB of switched systems with time‐varying delays via sampled‐data control are proposed. Moreover, considering the relationship between the sampling period and the mode‐dependent average dwell time, switching signals are designed. In addition, finite‐time weighted L₂‐gain (FTW‐L₂‐gain) of switched systems with time‐varying delays is proposed to measure their disturbance tolerance capacity within a finite‐time interval. Multiple Lyapunov‐Krasovskii functionals are applied to complete subsequent proofs in detail. Simulation results are exemplified to verify the proposed method.     

A one‐step leapfrog HIE‐FDTD method for rotationally symmetric structures

In this work, they propose a one‐step leapfrog hybrid implicit‐explicit finite‐difference time‐domain (HIE‐FDTD) method for body‐of‐revolution (BOR). Meanwhile, its Convolutional Perfect Matched Layer (CPML) absorbing boundary condition is implemented. In this method, the implicit difference is applied in the angular direction. All the resultant updating equations are still explicit. However, the stability condition of the proposed method is relaxed. The analytical analysis shows that its time step is only determined by the smaller one of spatial increments Δρ and Δz. A scattering example is provided to demonstrate the new algorithm. At the same time, the relative of reflection error of the CPML is given with comparisons of Mur.     

A 1‐to‐n way single‐ended‐to‐balanced substrate integrated waveguide filtering power splitter

An approach to 1‐to‐n (n = 3, 4…) way single‐ended‐to‐balanced filtering power splitter (SETBFPS) is proposed. The properly placed balanced ports with 0.5λ_g (λ_g is the substrate integrated waveguide [SIW] guided wavelength at f₀) space make the TE32nd 103 and TE32nd 105 modes of n 32nd‐mode SIW multimode resonators form differential‐mode (DM) passband of the SETBFPS. Compared with the state‐of‐art single‐ended‐to‐balanced power splitters, the proposed approach has all the functions of 1‐to‐n way, filtering, and common‐mode (CM) suppression. A 1‐to‐3 way prototype is exemplified at 3.5 GHz with the minimum insertion loss (IL) of 0.09 dB, a fractional bandwidth (FBW) for a 15‐dB return loss of 35%, and a FBW for 15‐dB CM suppression of 52%. Low IL and wide bandwidth can be observed.     

Composite disturbance‐observer‐based control and H ∞ control for nonlinear time‐delay systems

A novel type of control scheme combined the distance‐observer‐based control (DOBC) with H_∞ control is proposed for a class of nonlinear time‐delay systems subject to disturbances. The disturbances are supposed to include two parts. One in the input channel is generated by an exogenous system with uncertainty, which can represent the harmonic signals with modeling perturbations. The other is supposed to have the bounded H₂ norm. The disturbance observers based on regional pole placement and D‐stability theory are presented, which can be designed separately from the controller design. By integrating disturbance‐observer‐based control with H_∞ control laws, the disturbances can be rejected and attenuated, simultaneously, the desired dynamic performances can be guaranteed for nonlinear time‐delay systems with unknown nonlinear dynamics. Copyright © 2009 John Wiley and Sons Asia Pte Ltd and Chinese Automatic Control Society