scholarly journals Analysis of Serum Paraoxonase 1 Using Mass Spectrometry and Lectin Immunoassay in Patients With Alpha-Fetoprotein Negative Hepatocellular Carcinoma

2021 ◽  
Vol 11 ◽  
Author(s):  
Xinyi Cao ◽  
Zhao Cao ◽  
Yuyin Shao ◽  
Chao Liu ◽  
Guoquan Yan ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Hepatocellular Carcinoma ◽  
Inflammatory Diseases ◽  
Alpha Fetoprotein ◽  
Paraoxonase 1 ◽  
Label Free ◽  
The Difference ◽  
Validation Set ◽  
Serum Paraoxonase ◽  
Free Quantification

The diagnosis of AFP (alpha-fetoprotein)-negative HCC (hepatocellular carcinoma) mostly relies on imaging and pathological examinations, and it lacks valuable and practical markers. Protein N-glycosylation is a crucial post-translation modifying process related to many biological functions in an organism. Alteration of N-glycosylation correlates with inflammatory diseases and infectious diseases including hepatocellular carcinoma. Here, serum N-linked intact glycopeptides with molecular weight (MW) of 40–55 kDa were analyzed in a discovery set (n = 40) including AFP-negative HCC and liver cirrhosis (LC) patients using label-free quantification methodology. Quantitative lens culinaris agglutin (LCA) ELISA was further used to confirm the difference of glycosylation on serum PON1 in liver diseases (n = 56). Then, the alteration of site-specific intact N-glycopeptides of PON1 was comprehensively assessed by using Immunoprecipitation (IP) and mass spectrometry based 16O/18O C-terminal labeling quantification method to distinguish AFP-negative HCC from LC patients in a validation set (n = 64). Totally 195 glycopeptides were identified using a dedicated search engine pGlyco. Among them, glycopeptides from APOH, HPT/HPTR, and PON1 were significantly changed in AFP-negative HCC as compared to LC. In addition, the reactivity of PON1 with LCA in HCC patients with negative AFP was significantly elevated than that in cirrhosis patients. The two glycopeptides HAN253WTLTPLK (H5N4S2) and (H5N4S1) corresponding to PON1 were significantly increased in AFP-negative HCC patients, as compared with LC patients. Variations in PON1 glycosylation may be associated with AFP-negative HCC and might be helpful to serve as potential glycomic-based biomarkers to distinguish AFP-negative HCC from cirrhosis.

scholarly journals Label-Free Mass Spectrometry-Based Quantification of Linker Histone H1 Variants in Clinical Samples

2020 ◽  
Vol 21 (19) ◽  
pp. 7330
Author(s):  
Roberta Noberini ◽  
Cristina Morales Torres ◽  
Evelyn Oliva Savoia ◽  
Stefania Brandini ◽  
Maria Giovanna Jodice ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Histone H1 ◽  
Histone Variants ◽  
Linker Histone ◽  
Clinical Samples ◽  
Label Free ◽  
Complex Samples ◽  
Linker Histone H1 ◽  
Ideal Tool ◽  
Free Quantification

Epigenetic aberrations have been recognized as important contributors to cancer onset and development, and increasing evidence suggests that linker histone H1 variants may serve as biomarkers useful for patient stratification, as well as play an important role as drivers in cancer. Although traditionally histone H1 levels have been studied using antibody-based methods and RNA expression, these approaches suffer from limitations. Mass spectrometry (MS)-based proteomics represents the ideal tool to accurately quantify relative changes in protein abundance within complex samples. In this study, we used a label-free quantification approach to simultaneously analyze all somatic histone H1 variants in clinical samples and verified its applicability to laser micro-dissected tissue areas containing as low as 1000 cells. We then applied it to breast cancer patient samples, identifying differences in linker histone variants patters in primary triple-negative breast tumors with and without relapse after chemotherapy. This study highlights how label-free quantitation by MS is a valuable option to accurately quantitate histone H1 levels in different types of clinical samples, including very low-abundance patient tissues.

scholarly journals Fast Proteome Identification and Quantification from Data-Dependent Acquisition–Tandem Mass Spectrometry (DDA MS/MS) Using Free Software Tools

2019 ◽  
Vol 2 (1) ◽  
pp. 8 ◽  
Author(s):  
Jesse Meyer
Keyword(s):  
Mass Spectrometry ◽  
Tandem Mass Spectrometry ◽  
Gene Knockout ◽  
Tandem Mass ◽  
Label Free ◽  
Software Packages ◽  
Complex Process ◽  
Data Files ◽  
Using Data ◽  
Free Quantification

The identification of nearly all proteins in a biological system using data-dependent acquisition (DDA) tandem mass spectrometry has become routine for organisms with relatively small genomes such as bacteria and yeast. Still, the quantification of the identified proteins may be a complex process and often requires multiple different software packages. In this protocol, I describe a flexible strategy for the identification and label-free quantification of proteins from bottom-up proteomics experiments. This method can be used to quantify all the detectable proteins in any DDA dataset collected with high-resolution precursor scans and may be used to quantify proteome remodeling in response to drug treatment or a gene knockout. Notably, the method is statistically rigorous, uses the latest and fastest freely-available software, and the entire protocol can be completed in a few hours with a small number of data files from the analysis of yeast.

Label-Free Quantification by Data Independent Acquisition Mass Spectrometry to Map Cardiovascular Proteomes

2016 ◽  
pp. 227-245
Author(s):  
Sarah J. Parker ◽  
Ronald J. Holewinski ◽  
Irina Tchernyshyov ◽  
Vidya Venkatraman ◽  
Laurie Parker ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Label Free ◽  
Label Free Quantification ◽  
Data Independent Acquisition ◽  
Free Quantification

scholarly journals Host defense responses of CO441 and CL30 maize lines to Fusarium graminearum analyzed by comparative label-free quantitative proteomics

2019 ◽  
Author(s):  
Lana M. Reid ◽  
Illimar Altosaar
Keyword(s):  
Mass Spectrometry ◽  
Fusarium Graminearum ◽  
Fungal Pathogen ◽  
Detailed Knowledge ◽  
Label Free ◽  
Maize Line ◽  
Defense Proteins ◽  
Label Free Quantification ◽  
Host Pathogen ◽  
Free Quantification

AbstractGibberella ear rot is a disease of maize associated with low yields and the production of harmful mycotoxins therein. The disease is caused by the infection of host Zea mays with fungal pathogen Fusarium graminearum. Resistant (CO441) and susceptible (CL30) inbred maize line kernels were inoculated with conidial suspensions of F. graminearum or water (controls). Ears of maize (cobs) from each line were harvested upon maturation and proteins were extracted from the embryo tissue of the kernels to study tissue-specific response of the host. Embryo proteins from both CO441 and CL30 lines were sequenced using mass spectrometry (LC-MS/MS) and quantified using Label Free Quantification (LFQ). Following filtering, 509 proteins were identified. These proteins were grouped into nine functional categories: Fusarium-derived, late embryogenesis abundant, oil-body, metabolism, stress, cellular, protein storage, metabolism, and defense. Defense proteins were up-regulated in response to infection in both CO441 and CL30 lines. Furthermore, F. graminearum derived proteins were only found in CL30 infected kernels suggesting that resistance may be attributed in part to the inability of Fusarium to establish itself in the embryo. To our knowledge this is the first successful application of LFQ mass spectrometry to the study of host-pathogen response to F. graminearum.Biological significanceFungal pathogen Fusarium graminearum is responsible for billion dollar losses in crops and contamination of global grains with harmful mycotoxins. By studying host-pathogen interactions of Fusarium and maize on a proteomic level with resistant and susceptible genotypes, the biological interactions occurring during infection of the maturing seed were characterized. Mature kernels of the F. graminearum susceptible maize line CL30 and resistant CO441 line were dissected to permit a proteomic survey of the new sporophytic generation, the embryo. Detailed knowledge of this Host-pathogen interactome will assist development of new cereal lines resistant to the rot diseases caused by Fusarium graminearum.HighlightsSusceptible (CL30) and Resistant (CO441) lines were injected with water mock or F. graminearum LC-MS/MS of maize embryo protein extracts followed by Label Free Quantification (LFQ) permitted identification, quantification and comparison of proteomes between maize genotypes and treatments Fusarium-derived proteins were abundant only in the susceptible infected embryo Defense proteomes were up-regulated in both lines following infection nsLTP and Protease Inhibitor were significantly over-expressed in the Susceptible line after infection; chitinase and WIP1 were significantly over-expressed in the Resistant line after infection

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