scholarly journals A Shotgun Proteomic Platform for a Global Mapping of Lymphoblastoid Cells to Gain Insight into Nasu-Hakola Disease

2021 ◽  
Vol 22 (18) ◽  
pp. 9959
Author(s):  
Antonella De Palma ◽  
Anna Maria Agresta ◽  
Simona Viglio ◽  
Rossana Rossi ◽  
Maura D’Amato ◽  
...  
Keyword(s):  
Protein Identification ◽  
Case Reports ◽  
Protein Profile ◽  
Bone Lesions ◽  
Label Free ◽  
Lymphoblastoid Cells ◽  
Presenile Dementia ◽  
Signaling Complex ◽  
Global Mapping ◽  
On Line

Nasu-Hakola Disease (NHD) is a recessively inherited systemic leukodystrophy disorder characterized by a combination of frontotemporal presenile dementia and lytic bone lesions. NHD is known to be genetically related to a structural defect of TREM2 and DAP12, two genes that encode for different subunits of the membrane receptor signaling complex expressed by microglia and osteoclast cells. Because of its rarity, molecular or proteomic studies on this disorder are absent or scarce, only case reports based on neuropsychological and genetic tests being reported. In light of this, the aim of this paper is to provide evidence on the potential of a label-free proteomic platform based on the Multidimensional Protein Identification Technology (MudPIT), combined with in-house software and on-line bioinformatics tools, to characterize the protein expression trends and the most involved pathways in NHD. The application of this approach on the Lymphoblastoid cells from a family composed of individuals affected by NHD, healthy carriers and control subjects allowed for the identification of about 3000 distinct proteins within the three analyzed groups, among which proteins anomalous to each category were identified. Of note, several differentially expressed proteins were associated with neurodegenerative processes. Moreover, the protein networks highlighted some molecular pathways that may be involved in the onset or progression of this rare frontotemporal disorder. Therefore, this fully automated MudPIT platform which allowed, for the first time, the generation of the whole protein profile of Lymphoblastoid cells from Nasu-Hakola subjects, could be a valid approach for the investigation of similar neurodegenerative diseases.

scholarly journals Two-dimensional gel electrophoretic protein profile analysis during seed development of Ocotea catharinensis: a recalcitrant seed species

2010 ◽  
Vol 22 (1) ◽  
pp. 23-33 ◽  
Author(s):  
Leonardo L.C. Dias ◽  
Tiago S. Balbuena ◽  
Vanildo Silveira ◽  
Claudete Santa-Catarina ◽  
Andrej Schevchenko ◽  
...  
Keyword(s):  
Seed Development ◽  
Protein Identification ◽  
Protein Extraction ◽  
Profile Analysis ◽  
Protein Profile ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis ◽  
Recalcitrant Seed ◽  
And Storage ◽  
Cotyledonary Stage

The aim of the present work was to characterize changes in the protein profile throughout seed development in O. catharinensis, a recalcitrant species, by two-dimensional gel electrophoresis. Protein extraction was undertaken by using a thiourea/urea buffer, followed by a precipitation step with 10% TCA. Comparative analysis during seed development showed that a large number of proteins were exclusively detected in each developmental stage. The cotyledonary stage, which represents the transition phase between embryogenesis and the beginning of metabolism related to maturation, presents the highest number of stage-specific spots. Protein identification, through MS/MS analysis, resulted in the identification of proteins mainly related to oxidative metabolism and storage synthesis. These findings contribute to a better understanding of protein metabolism during seed development in recalcitrant seeds, besides providing information on established markers that could be useful in defining and improving somatic embryogenesis protocols, besides monitoring the development of somatic embryos in this species.

Multiple Myeloma Associated GI Polyposis

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 5009-5009
Author(s):  
Nassim Nabbout ◽  
Mohamad El Hawari ◽  
Thomas K. Schulz
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Plasma Cells ◽  
Conflicts Of Interest ◽  
Case Reports ◽  
Bone Marrow Failure ◽  
Bone Lesions ◽  
Rare Manifestation ◽  
History Of ◽  
Central Ulceration

Abstract Abstract 5009 Multiple myeloma is a neoplastic proliferation of monoclonal plasma cells that can result in osteolytic bone lesions, hypercalcemia, renal impairment, bone marrow failure, and the production of monoclonal gammopathy. The gastrointestinal tract is rarely involved in myeloma. GI polyposis is a rare manifestation of extra-medullary disease in multiple myeloma. Such cases usually present as gastrointestinal hemorrhage or intestinal obstruction. A 53-year-old African American male recently diagnosed with multiple myeloma presented with three-day history of rectal bleed and fatigue. EGD showed multiple raised, polypoid, rounded lesions with a superficial central ulceration in the stomach. Colonoscopy showed similar lesions in the ascending and transverse areas of the colon that ranged in size from 5 to 16 mm in diameter. Biopsies showed that these polyps were made of plasma cells. A bone marrow biopsy showed diffuse involvement (greater than 90%) of bone marrow with multiple myeloma with anaplastic features. The patient was started on bortezomib at diagnosis, however, he passed away a few weeks later. This type of metastatic disease has been described in isolated case reports in the literature, while solitary GI plasmacytoma has been reported more frequently. In rare cases, multiple myeloma can involve the GI tract which may lead to bleed or obstruction. This involvement is likely a marker of aggressivity. This example of extra-medullary disease in myeloma is an uncommon variant with features of poor prognosis and dedifferentiation. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Improved Label-Free LC-MS Analysis by Wavelet-Based Noise Rejection

2010 ◽  
Vol 2010 ◽  
pp. 1-9 ◽  
Author(s):  
Salvatore Cappadona ◽  
Paolo Nanni ◽  
Marco Benevento ◽  
Fredrik Levander ◽  
Piera Versura ◽  
...  
Keyword(s):  
Protein Identification ◽  
Label Free ◽  
Computational Framework ◽  
Noise Rejection ◽  
Software Packages ◽  
Ms Analysis ◽  
More Care ◽  
Multiple Samples ◽  
Common Laboratory

Label-free LC-MS analysis allows determining the differential expression level of proteins in multiple samples, without the use of stable isotopes. This technique is based on the direct comparison of multiple runs, obtained by continuous detection in MS mode. Only differentially expressed peptides are selected for further fragmentation, thus avoiding the bias toward abundant peptides typical of data-dependent tandem MS. The computational framework includes detection, alignment, normalization and matching of peaks across multiple sets, and several software packages are available to address these processing steps. Yet, more care should be taken to improve the quality of the LC-MS maps entering the pipeline, as this parameter severely affects the results of all downstream analyses. In this paper we show how the inclusion of a preprocessing step of background subtraction in a common laboratory pipeline can lead to an enhanced inclusion list of peptides selected for fragmentation and consequently to better protein identification.

On-line learning with minimized change of the global mapping

2014 ◽  
Vol 6 (2) ◽  
pp. 131-151
Author(s):  
Andreas Buschermöhle ◽  
Werner Brockmann
Keyword(s):  
Global Mapping ◽  
On Line ◽  
On Line Learning

scholarly journals Acoustic Immunosensing of Exosomes Using a Quartz Crystal Microbalance with Dissipation Monitoring

2019 ◽  
Author(s):  
Jugal Suthar ◽  
Edward Parsons ◽  
Bart Hoogenboom ◽  
Gareth Williams ◽  
Stefan Guldin
Keyword(s):  
Quartz Crystal Microbalance ◽  
Quartz Crystal ◽  
Lipid Membrane ◽  
Acoustic Analysis ◽  
Limit Of Detection ◽  
Culture Media ◽  
Transmembrane Protein ◽  
Protein Profile ◽  
Size Exclusion ◽  
Label Free

Exosomes are endocytic lipid-membrane bound bodies with potential to be used as biomarkers in cancer and neurodegenerative disease. The limitations and scarcity of current exosome characterisation approaches has led to a growing demand for translational techniques, capable of determining their molecular composition and physical properties in physiological fluids. Here, we investigate label-free immunosensing, using a quartz crystal microbalance with dissipation (QCM-D), to detect exosomes by exploiting their surface protein profile. Exosomes expressing the transmembrane protein CD63 were isolated by size-exclusion chromatography from cell culture media. QCM-D sensors functionalised with anti-CD63 antibodies formed a direct immunoassay towards CD63-positive exosomes, exhibiting a limit-of-detection of 1.7x10^8 and 1.1x10^8 exosome sized particles (ESPs) ml^-1 for frequency and dissipation response respectively, i.e., clinically relevant concentrations. Our proof-of-concept findings support the adoption of dual-mode acoustic analysis of exosomes, leveraging both frequency and dissipation monitoring for use in diagnostic assays.

scholarly journals The beauty of being (label)-free: sample preparation methods for SWATH-MS and next-generation targeted proteomics

F1000Research ◽  
2014 ◽  
Vol 2 ◽  
pp. 272 ◽  
Author(s):  
Jakob Vowinckel ◽  
Floriana Capuano ◽  
Kate Campbell ◽  
Michael J. Deery ◽  
Kathryn S. Lilley ◽  
...  
Keyword(s):  
Sample Preparation ◽  
Protein Identification ◽  
Protein Detection ◽  
Cost Effective ◽  
High Signal ◽  
Label Free ◽  
Signal Variation ◽  
Signal Stability ◽  
Advantages And Disadvantages ◽  
Free Data

The combination of qualitative analysis with label-free quantification has greatly facilitated the throughput and flexibility of novel proteomic techniques. However, such methods rely heavily on robust and reproducible sample preparation procedures. Here, we benchmark a selection of in gel, on filter, and in solution digestion workflows for their application in label-free proteomics. Each procedure was associated with differing advantages and disadvantages. The in gel methods interrogated were cost effective, but were limited in throughput and digest efficiency. Filter-aided sample preparations facilitated reasonable processing times and yielded a balanced representation of membrane proteins, but led to a high signal variation in quantification experiments. Two in solution digest protocols, however, gave optimal performance for label-free proteomics. A protocol based on the detergent RapiGest led to the highest number of detected proteins at second-best signal stability, while a protocol based on acetonitrile-digestion, RapidACN, scored best in throughput and signal stability but came second in protein identification. In addition, we compared label-free data dependent (DDA) and data independent (SWATH) acquisition on a TripleTOF 5600 instrument. While largely similar in protein detection, SWATH outperformed DDA in quantification, reducing signal variation and markedly increasing the number of precisely quantified peptides.

scholarly journals Software for Gene-Biosensor based on PCR and Electrical Bioimpedance

2021 ◽  
Vol 2008 (1) ◽  
pp. 012014
Author(s):  
I A Castillo-Salazar ◽  
G Ames-Lastra ◽  
E Sacristán-Rock ◽  
A Hernández-Nava ◽  
C A González-Díaz
Keyword(s):  
Control Program ◽  
Integrated System ◽  
Label Free ◽  
Development Environment ◽  
Language Version ◽  
Python Language ◽  
Pcr Product ◽  
Multifrequency Bioimpedance ◽  
Electrical Bioimpedance ◽  
On Line

Abstract Gene detection by the use of bioimpedance measurements is an emerging technical proposal from the last two decades. Our recent studies have shown the feasibility to use multifrequency bioimpedance to detect specific label-free Deoxyribonucleic Acids (DNA) sequence in the final product of Polymerase Chain Reaction (PCR). We have developed a gene-biosensor integrated by a thermocycler block for final point PCR process and a relative bioimpedance meter at every PCR cycle in the sample. The system demands a dynamic software for all gene-biosensor modules control. This work reports a control program design for a gene-biosensor based on PCR product and bioimpedance measurements, the general structural philosophy, operating routines and subroutines as well as its interaction with the hardware in every module are described. The program was designed on the basis of Python language version 3.8.3 with the support of the Visual Studio Code as Integrated Development Environment (IDE), and using Windows 10 as the operating system. Results indicate the control program allows a suitable governing of the bioimpedance meter and PCR thermocycler block, both as a well-integrated system. Bioimpedance and temperature measurements are in agreement with the control operating structural design. Additional amendments regarding an on-line monitoring system are warranted.

scholarly journals Development of a NanoMIPs-SPR-Based Sensor for β-Lactoglobulin Detection

Chemosensors ◽  
2020 ◽  
Vol 8 (4) ◽  
pp. 94
Author(s):  
Roberta D’Aurelio ◽  
Jon Ashley ◽  
Thomas L. Rodgers ◽  
Linda Trinh ◽  
Jeff Temblay ◽  
...  
Keyword(s):  
Solid Phase ◽  
Cross Contamination ◽  
Label Free ◽  
Phase Synthesis ◽  
Processing Plants ◽  
On Line ◽  
Amine Groups ◽  
Final Optimization ◽  
Β Lactoglobulin ◽  
Selective Label

Food manufacturers are aiming to manage the levels of cross-contamination of allergens within food processing plants and ultimately move away from precautionary labelling. Hence, the need for rapid methods to detect allergens cross-contamination. A sensitive and selective label-free nanoMIPs based sensor was developed and tested for the detection of β-lactoglobulin (BLG). NanoMIPs were synthesized using solid-phase synthesis and appeared as spherical nanoparticles with sizes ranging from 264–294 nm, using dynamic light scattering (DLS). The nanoMIPs were functionalized with amine groups and attached to the surface of the SPR gold chip via amine-coupling protocol. The SPR nanoMIPs-based sensor demonstrated a detection limit of 3 ng mL−1 (211 pM) over a linear range of 1–5000 ng mL−1, with binding affinity of 7.0 × 10−8 M and specificity towards BLG. With further testing and final optimization, the developed nanosensor can be integrated on-line or at-line cleaning-in-place (CIP) wash systems, allowing to effectively monitor milk protein allergens as a rapid, point-of-source methodology.

Proper imputation of missing values in proteomics datasets for differential expression analysis

2020 ◽  
Author(s):  
Mingyi Liu ◽  
Ashok Dongre
Keyword(s):  
Differential Expression ◽  
Expression Analysis ◽  
Protein Identification ◽  
Missing Values ◽  
Tissue Sample ◽  
Differential Expression Analysis ◽  
Real Life ◽  
Label Free ◽  
Imputation Methods ◽  
The Impact

Abstract Label-free shotgun proteomics is an important tool in biomedical research, where tandem mass spectrometry with data-dependent acquisition (DDA) is frequently used for protein identification and quantification. However, the DDA datasets contain a significant number of missing values (MVs) that severely hinders proper analysis. Existing literature suggests that different imputation methods should be used for the two types of MVs: missing completely at random or missing not at random. However, the simulated or biased datasets utilized by most of such studies offer few clues about the composition and thus proper imputation of MVs in real-life proteomic datasets. Moreover, the impact of imputation methods on downstream differential expression analysis—a critical goal for many biomedical projects—is largely undetermined. In this study, we investigated public DDA datasets of various tissue/sample types to determine the composition of MVs in them. We then developed simulated datasets that imitate the MV profile of real-life datasets. Using such datasets, we compared the impact of various popular imputation methods on the analysis of differentially expressed proteins. Finally, we make recommendations on which imputation method(s) to use for proteomic data beyond just DDA datasets.

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