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Results for “differential proteomics analysis”

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PubMed2006

2-DE proteomic profiling of neuronal stem cells.

Proteomics has become a powerful tool in neuroscience studies. Although numerous human neural stem cells are available for research purposes since many years, there exists only limited information on proteomic data from stable neural stem cell lines. Profiling and functional proteome studies of neuronal stem cells will help to describe the protein inventory as well as protein activity and interactions, subcellular localization and posttranslational modifications. The proteomic analysis of neuronal differentiation processes will elucidate the complex events leading to the generation of different phenotypes via distinctive developmental programs that control self-renewal, differentiation, and plasticity. Using the ReNcell VM197 model, a cell line derived from human fetal ventral mesencephalon stem cells, we studied the protein inventory of the stem cells by 2-DE gel electrophoresis and mass spectrometric protein identification and constructed a 2-DE protein map consisting of more than 400 identified protein spots. This proteome reference database constitutes the basis for further investigations of differential protein expression during differentiation. A profiling of the neuronal differentiation-associated changes displayed the large rearrangement of the proteome during this process, and the proteomic techniques proved to be a valuable tool for the elucidation of neuronal differentiation process and for target protein screening.

Animals↗

PubMed2006

Proteomic search for potential diagnostic markers and therapeutic targets for ovarian clear cell adenocarcinoma.

Clear cell adenocarcinoma (CCA) has a highly malignant potential in human epithelial ovarian cancer. The serum CA-125 is widely used as a marker for ovarian cancer, but the level is relatively low in CCA. Therefore, new sensitive biomarkers are required. In this report, we describe a promising proteomic analysis that is differentially expressed in CCA when compared to mucinous adenocarcinoma, using the ovarian cultured cell lines OVISE, OVTOKO, and MCAS. The disease-associated proteins were identified by 2-D differential gel electrophoresis (2-D DIGE) and MS. In this analysis, 18 up-regulated and 31 down-regulated spots were observed that had at least two-fold differences in the two CCA cell lines than in MCAS as control cells. Some of the proteins differentially expressed in CCA were previously observed as alternative expression levels in ovarian and/or other cancers in clinical samples. In a subsequent preliminary differential study using surgical specimens from patients with CCA, it was demonstrated that the identified proteins were expressed differentially in actual tissues, as well as in the CCA culture cells. The results from this investigation show the potentiality of a proteomic approach for identifying disease-associated proteins, which may eventually serve as diagnostic markers or therapeutic targets in CCA.

Adenocarcinoma, Clear Cell↗

PubMed2026

Multi-Omics Analysis Reveals Molecular Networks and Key Pathways Associated with Cysteine- and Methionine-Mediated Biosynthesis of Sulfur-Containing Flavor Metabolites in Lentinula edodes.

Lentinula edodes is renowned for its unique aroma, which is characterized by various volatile sulfur-containing flavor metabolites (SCFMs). Cysteine and methionine could enhance the SCFMs biosynthesis in L. edodes; however, the underlying metabolic pathways remain unclear. To bridge this gap, integrated proteomic and metabolomic analysis were performed to decipher pathways through which cysteine and methionine regulate SCFM biosynthesis. Results showed that exogenous cysteine and methionine supplementation significantly increased the content of lenthionine, the key aroma compound of shiitake mushrooms. Both treatments induced substantial changes in the proteomic and metabolomic profiles. Proteomic analysis revealed that differentially expressed proteins were predominantly enriched in cysteine and methionine metabolism and sulfur metabolism following cysteine treatment, whereas methionine treatment mainly affected proteins associated with tryptophan metabolism and sulfur metabolism. Metabolomic analysis showed that differentially accumulated metabolites were significantly enriched in D-amino acid metabolism and cysteine and methionine metabolism, with glutathione metabolism specifically enriched under cysteine treatment. Integrated omics analysis further uncovered distinct sulfur metabolite-protein regulatory networks under different sulfur nutrition and identified treatment-specific hub proteins. These findings establish a molecular regulatory framework linking SCFM biosynthesis with broader primary metabolic pathways involved in sulfur intermediate generation and regulation, providing new insights into the potential regulatory networks underlying SCFM formation in L. edodes.

Methionine↗

PubMed2026

Proteomic Analysis of Extracellular Vesicles Reveals Vitronectin and Laminin Subunit Alpha-3 as Candidate Biomarkers for Gastric Cancer.

BACKGROUND/AIMS: Clinically useful noninvasive biomarkers for gastric cancer remain limited. Extracellular vesicles (EVs) carry a molecular cargo reflective of their cells of origin and have emerged as promising candidates for blood-based cancer biomarkers. We aimed to identify EV-associated protein biomarkers for gastric cancer via a proteomic approach. METHODS: Proteomic profiling of EVs was performed using one normal gastric cell line (Hs738st/int) and two gastric cancer cell lines (AGS and NCI-N87). Selected proteins were validated in blood-derived EVs isolated from plasma samples of 10 healthy controls and 36 patients with gastric cancer. RESULTS: Proteomic analysis identified 224 differentially expressed proteins whose expression was consistently altered in gastric cancer cell line-derived EVs. Among these, vitronectin (VTN) and laminin subunit alpha-3 (LAMA3) were selected based on their consistent upregulation. EV-associated LAMA3 levels were significantly higher in patients with gastric cancer than in healthy controls (p=0.003), with significant elevations observed from stage II onward (p=0.041, p=0.017, and p=0.004 for stages II, III, and IV, respectively). EV-associated VTN levels were not significantly different overall (p=0.089); however, stage-specific analysis demonstrated significant increases in VTN levels in patients with stage III (p=0.036) and stage IV (p=0.005) gastric cancer. Both EV-associated VTN and LAMA3 levels showed significant positive correlations with the cancer stage (ρ=0.564 and ρ=0.611, respectively; both p<0.001). CONCLUSIONS: The levels of EV-associated VTN and LAMA3 appear to be more closely associated with disease progression than with early-stage detection of gastric cancer. These findings suggest that EV-based proteomic biomarkers may have clinical utility for monitoring tumor progression in patients with clinically advanced gastric cancer.

Humans↗

PubMed2026

11-O-galloylbergenin alleviates LPS-stimulated inflammation in RAW 264.7 macrophages by targeting Grb2, RhoA, and Cdc42 in the RAS signaling pathway.

OBJECTIVE: This study aimed to explore the anti-inflammatory mechanism of 11-O-galloylbergenin in macrophages. METHODS: Lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages were treated with 11-O-galloylbergenin. Cytotoxicity was assessed by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, and cytokine secretion was measured by enzyme-linked immunosorbent (ELISA) assay. Data-independent acquisition (DIA)-based proteomics, Gene Ontology (GO)/Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, western blotting, and molecular docking were also performed. RESULTS: 11-O-Galloylbergenin (≤50 μg/mL) was non-cytotoxic and significantly suppressed tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) secretion. Proteomics analysis identified 209 differentially expressed proteins (DEPs) that showed opposite expression trends before and after 11-O-galloylbergenin treatment. Seven of these DEPs were enriched in the RAS pathway: RhoA, Cdc42, Grb2, RalB, Calm3, Gnb2, and Pla2g4a. Western blotting confirmed that 11-O-galloylbergenin downregulated RhoA, Cdc42, and Grb2 expression. Molecular docking revealed good binding affinity of 11-O-galloylbergenin to RhoA, Cdc42, and Grb2. CONCLUSION: 11-O-Galloylbergenin alleviates LPS-stimulated inflammation in RAW 264.7 macrophages by inhibiting the RAS signaling pathway.

Animals↗

PubMed2004

Proteomic analysis of glutathione S -transferases of Arabidopsis thaliana reveals differential salicylic acid-induced expression of the plant-specific phi and tau classes.

Plant glutathione S -transferases (GSTs) are a large group of multifunctional proteins that are induced by diverse stimuli. Using proteomic approaches we identified 20 GSTs at the protein level in Arabidopsis cell culture with a combination of GST antibody detection, LC-MS/MS analysis of 23-30 kDa proteins and glutathione-affinity chromatography. GSTs identified were from phi, tau, theta, zeta and DHAR sub-sections of the GST superfamily of 53 members. We have uncovered preliminary evidence for post-translational modifications of plant GSTs and show that phosphorylation is unlikely to be responsible. Detailed analysis of GST expression in response to treatment with 0.01-1 mM of the plant defence signal salicylic acid (SA) uncovered some interesting features. Firstly, GSTs appear to display class-specific concentration-dependent SA induction profiles highlighting differences between the large, plant specific phi and tau classes. Secondly, different members of the same class, while sharing similar SA dose responses, may display differences in terms of magnitude and timing of induction, further highlighting the breadth of GST gene regulation. Thirdly, closely related members of the same class ( GSTF6 and GSTF7 ), arising via tandem duplication, may be regulated differently in terms of basal expression levels and also magnitude of induction raising questions about the role of subfunctionalisation within this family. Our results reveal that GSTs exhibit class specific responses to SA treatment suggesting that several mechanisms are acting to induce GSTs upon SA treatment and hinting at class-specific functions for this large and important, yet still relatively elusive gene family.

Arabidopsis↗

PubMed2006

Semi-supervised LC/MS alignment for differential proteomics.

MOTIVATION: Mass spectrometry (MS) combined with high-performance liquid chromatography (LC) has received considerable attention for high-throughput analysis of proteomes. Isotopic labeling techniques such as ICAT [5,6] have been successfully applied to derive differential quantitative information for two protein samples, however at the price of significantly increased complexity of the experimental setup. To overcome these limitations, we consider a label-free setting where correspondences between elements of two samples have to be established prior to the comparative analysis. The alignment between samples is achieved by nonlinear robust ridge regression. The correspondence estimates are guided in a semi-supervised fashion by prior information which is derived from sequenced tandem mass spectra. RESULTS: The semi-supervised method for finding correspondences was successfully applied to aligning highly complex protein samples, even if they exhibit large variations due to different biological conditions. A large-scale experiment clearly demonstrates that the proposed method bridges the gap between statistical data analysis and label-free quantitative differential proteomics. AVAILABILITY: The software will be available on the website http://people.inf.ethz.ch/befische/proteomics.

Algorithms↗

PubMed2025

Protective effects of liver-derived apolipoprotein A1 against heat stress-induced hypothalamic lipid metabolism and blood-brain barrier integrity.

Heat stress (HS), a prevalent occupational and environmental hazard, has increasingly been recognized as a major contributor to multiple physiological disorders. The hypothalamus, a key regulator of thermoregulation and endocrine signaling, is especially susceptible to metabolic and inflammatory disturbances induced by HS. This study investigates the interplay among lipid metabolism, blood-brain barrier (BBB) integrity, and neuroinflammation in the hypothalamus under HS conditions, with a specific focus on apolipoprotein A1 (APOA1) as a potential protective factor. To achieve this, we integrated proteomic and lipidomic analyses with experimental validation in porcine and murine models. Proteomic analysis identified 266 differentially expressed proteins (DEPs) in the hypothalamus following HS, with significant enrichment in lipid metabolism pathways-especially glycerophospholipid (GP) metabolism-in which APOA1 displayed a marked increase. Lipidomic profiling further revealed HS-induced disruptions in phosphatidylcholine (PC), phosphatidylethanolamine (PE), and cardiolipin (CL) metabolism. Additionally, blood-brain barrier integrity was compromised, as evidenced by increased perivascular IgG extravasation, reduced pericyte coverage, and decreased expression of tight junction proteins ZO-1 and Occludin. HS also triggered pronounced neuroinflammation, characterized by elevated levels of iNOS, GFAP, and pro-inflammatory cytokines (TNF-α, IL-1β, and IL-6). Notably, administration of D-4F, an APOA1 mimetic peptide, alleviated blood-brain barrier damage, reduced neuroinflammation, and preserved synaptic integrity, thereby suggesting a neuroprotective role for APOA1 in HS-induced hypothalamic dysfunction. These findings underscore the critical role of lipid metabolism in maintaining hypothalamic homeostasis under HS conditions and position APOA1 as a key regulator with potential therapeutic implications for mitigating HS-related neuroinflammatory and metabolic disturbances.

Blood-Brain Barrier↗

PubMed2006

Considerations for proteolytic labeling-optimization of 18O incorporation and prohibition of back-exchange.

Proteolytic (18)O labeling is a very powerful tool for differential analysis applied to proteome studies. However, it is a relatively new technique and the optimization of the labeling process still needs some attention. We found that the two-step post-proteolytic labeling should be favored over the conventional digestion of proteins in H(2) (18)O, since the former allows for higher sample concentrations and thus more favorable kinetics. It was demonstrated that the inhibitory effect of urea on (18)O incorporation could be compensated by the use of higher sample concentrations. Furthermore, it was shown that heat-deactivation of trypsin prevents (18)O/(16)O back-exchange. In addition, no non-specific hydrolysis of the peptides could be observed as a result of the heating. Heat inactivation of trypsin opens the way for the use of capillary electrophoresis as a separation technique in proteolytic labeling studies, as it abolishes the need for use of detrimental additives. Analysis of a labeled protein digest by capillary isoelectric focusing/mass spectrometry showed the applicability of the method. No back-exchange was observed across the entire electropherogram.

Electrophoresis, Capillary↗

PubMed2025

Gain-of-function PPM1D mutations attenuate ischemic stroke.

Identification of genetic aberrations in stroke, the second leading cause of death worldwide, is of paramount importance for understanding the disease pathogenesis and generating new therapies. Whole-genome sequencing from 10,241 ischemic stroke patients identified eight patients carrying gain-of-function mutations on coding variants in the protein phosphatase magnesium-dependent 1 δ (PPM1D) gene. Patients carrying PPM1D mutations exhibit better stroke-related clinical phenotypes, including improvements in peripheral inflammation, fibrinogen, low-density lipoprotein, cholesterol and plateletcrit level. Experimental brain ischemia in Ppm1d-deficient (Ppm1d-/-) mice resulted in enlarged lesions and pronounced neurological impairments. Spatial transcriptomics revealed a distinct Ppm1d-associated gene expression pattern, indicating disrupted endothelial homeostasis during ischemic brain injury. Proteomic analysis demonstrated that differentially expressed proteins in primary brain endothelial cells from Ppm1d-/- mice were significantly enriched in the peroxisome proliferator-activated receptors (PPARs)-mediated metabolic signaling. Mechanistically, Ppm1d deficiency promoted aberrant fatty acid β-oxidation and increased oxidative stress, which impaired endothelial cell function through the PPARα pathway. A small molecule, T2755, was identified to engage Trp427 and stabilize PPM1D, thereby mitigating ischemic brain injury in mice. Collectively, we find that PPM1D protects against ischemic brain injury and validates its pharmacological stabilizer T2755 as a promising therapy for ischemic stroke. Gain-of-function PPM1D mutations attenuate ischemic cerebral injury. Whole-genome sequencing data of 10,241 ischemic stroke patients from the Third Chinese National Stroke Registry (CNSR-III) identified eight patients with gain-of-function mutations in the protein phosphatase magnesium-dependent 1 δ (PPM1D) gene (17q23.2). These mutation carriers displayed improved peripheral inflammation, decreased fibrinogen, low-density lipoprotein, cholesterol and plateletcrit level. Ppm1d-deficient (Ppm1d-/-) mice exhibited exacerbated stroke outcomes, characterized by enlarged infarct volumes, disrupted cerebrovascular architecture, and enhanced neuro-inflammation. Mechanistically, Ppm1d deficiency induced the disturbance of endothelial fatty acid metabolism involving the PPARα pathway. Through integrated computational modeling, virtual screening, and in vitro validation, T2755 was identified as a small molecule PPM1D stabilizer. Pharmacological PPM1D stabilization with T2755 significantly attenuated ischemic brain injury in murine models.

Aged↗

PubMed2026

Hepatic metabolic adaptation to endurance exercise: temporal and sex differences by multiomics integration and validation.

BACKGROUND: Although endurance exercise benefits liver health, sex-specific adaptive trajectories remain unclear. This study mapped dynamic liver adaptation in males and females during prolonged training and identified underlying molecular programs. METHODS: Using publicly available time-resolved liver multi-omics data generated by the Molecular Transducers of Physical Activity Consortium (MoTrPAC), we established a computational pipeline for differential analysis of transcriptomic, proteomic, phosphoproteomic, and metabolomic data with FDR correction, followed by FGSEA pathway enrichment. Kinase activities were inferred through ortholog mapping and PhosphoSitePlus. Cross-omics co-expression networks were constructed using WGCNA and topological overlap to link omics features with physiological phenotypes. For experimental validation, liver tissues were collected from endurance-trained Sprague-Dawley rats, and key nodes were confirmed by Western blotting, qRT-PCR, and immunofluorescence/immunohistochemical staining. Public scRNA-seq data were further integrated to map multi-omics signals to single-cell resolution and assess functional changes in specific cell types. RESULTS: The hepatic response to exercise stress was stage-specific, shifting from early transcriptional activation to later proteomic and metabolic remodeling. Multi-omics integration revealed distinct sex-associated adaptive trajectories: males were more strongly associated with energy metabolism, redox-related programs, and amino acid/organic acid catabolism, whereas females showed prominent membrane lipid remodeling, proteostasis -related programs, and mitochondrial/ribosomal translational features. Single-cell analysis showed that tissue remodeling occurred without major lineage turnover, instead involving altered communication among pre-existing cell communities. Validation of PPP1R3G identified a protein-dominant exercise-responsive marker, supporting the contribution of post-transcriptional or protein-level regulation. CONCLUSIONS: Hepatic adaptation to endurance stress follows a cross-omics evolutionary pattern with sex-specific reprogramming of energy supply and homeostatic maintenance. This time-resolved framework clarifies how exercise improves liver function and supports sex-oriented metabolic interventions and therapeutic target discovery.

Animals↗

PubMed2004

Identification of tumor markers using two-dimensional electrophoresis in gastric carcinoma.

AIM: To study the differential expression of proteins in normal and cancerous gastric tissues, and further identify new molecular markers for diagnosis and prognosis of gastric carcinoma, as well as develop new therapeutic targets of the disease. METHODS: Matched pairs of tissues from 6 gastric cancer patients were analyzed for their two-dimensional electrophoresis (2DE) profiles. Soluble fraction proteins from human normal and cancerous gastric tissue were separated in the first dimension by isoelectric focusing on immobilized pH gradient (IPG, pH3-10) strips, and by 125 g/L sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) in the second dimension with silver nitrate staining. Protein differential expression was analyzed by use of image analysis software to find out candidates for gastric cancer-associated proteins. RESULTS: Nine protein spots overexpressed in tumor tissues as compared with noncancerous regions. In the next step, 9 tumor-specific spots were cut off from Coomassie Brilliant Blue staining gels, digested in gel with L-1-tosylamide-2-phenylethyl chloromethyl ketone (TPCK)-trypsin. Protein identification was done by peptide mass fingerprinting with matrix assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS). In total, 5 tumor-specific protein spots corresponding to 5 different polypeptide chains were identified, including annexin V, carbonic anhydrase, prohibitin, fibrin beta and fibrinogen fragment D. Among these 5 spots, the potential significance of the differential expressions is discussed. CONCLUSION Differential expression analysis of proteomes may be useful for the development of new molecular markers for diagnosis and prognosis of gastric carcinoma.

Biomarkers, Tumor↗

PubMed2006

A systems approach to model metastatic progression.

Proteomic profiling of human disease has seen much early activity with the accessibility of the newest generation of high-throughput platforms and technologies. Nevertheless, the nature of the dynamic physiologic milieu and high dimensionality of the data has complicated major diagnostic and prognostic breakthroughs. Our recent article in Cancer Cell delineates an integrative model for culling a molecular signature of metastatic progression in prostate cancer from proteomic and transcriptomic analyses and shows its facility as a predictor of prognosis. The study leveraged direct proteomic analysis of tumor tissue extracts, differential feature selection characterizing the proteomic alterations of prostate cancer subclasses, and integration with public and study-derived genomic data to construct a multiplex gene signature representing progression of indolent cancer to aggressive disease. This further predicted clinical outcome in a variety of solid tumors. This review describes the context of the work, the framework for the analysis itself, and a look forward to the promise of this systems approach to human disease.

Computational Biology↗

PubMedDate not supplied

Proteomic analysis of the genetic premature aging disease Hutchinson Gilford progeria syndrome reveals differential protein expression and glycosylation.

Proteomics has revealed differential protein expression and glycosylation in membrane proteins from premature aging Hutchinson-Gilford progeria syndrome fibroblasts (progeria). Progeria is a rare autosomal dominant genetic disorder of premature aging characterized by marked growth retardation and specific, progressive, premature senescent changes of the skin and other tissues. Affected children live to an average age of 13 years. The 1q20-24 region of chromosome 1 which codes for one of these proteins, lamin A/C, has previously been implicated by Brown et al. (1990) who described identical twins with progeria, where cytogenetic analysis showed an inverted insertion in the long arm of the chromosome in 70% of cells. Luengo et al. (2002) similarly reported an interstitial deletion of chromosome 1q23, in a 9-year-old patient with a classic clinical picture of progeria.

Aging, Premature↗

PubMed2004

A proteomic analysis identifies glutathione S-transferase isoforms whose abundance is differentially regulated by ethylene during the formation of early root epidermis in Arabidopsis seedlings.

The plant hormone ethylene has been shown to play an important role in root hair development in Arabidopsis. With the aid of proteomic analysis, we identified three distinct glutathione S-transferase (GST) isoforms, AtGSTF2, AtGSTF8, and AtGSTU19, expressed early in root epidermal establishment in Arabidopsis seedlings. The AtGSTF2 protein was specifically up-regulated by ethylene. A subsequent RNA expression study revealed that the AtGSTF2 gene was highly sensitive to ethylene, whereas the transcripts for AtGSTF8 and AtGSTU19 were constitutively present in new root tissue of 4-day-old seedlings. The steady-state level of AtGSTF2 mRNA was greatly reduced in the roots of ethylene-insensitive mutants, while mutation at the CTR1 locus, which confers an ectopic root hair phenotype, resulted in a markedly elevated level of AtGSTF2 transcript in young root tissue. Although the physiological function of ethylene-induced AtGSTF2 is not yet clear, there are several possibilities for its role during early root development.

Amino Acids, Cyclic↗

PubMed2004

Comparative analysis of phosphoprotein-enriched myocyte proteomes reveals widespread alterations during differentiation.

The differentiation of skeletal muscle has been associated with altered phosphorylation status of individual proteins. However, a global analysis of protein phosphorylation during myogenesis has yet to be undertaken. Here, we report the identification of over 130 putative phosphoproteins from murine C2C12 muscle cells. Cell extracts were fractionated on phosphoprotein enrichment columns and the resulting proteins were detected by two-dimensional gel electrophoresis and silver stain, and identified by liquid chromatography coupled to electrospray tandem mass spectrometry. The early differentiation of C2C12 myoblasts was found to be accompanied by changes in the phosphorylation or expression of numerous proteins including cytoskeletal, heat shock and signaling proteins, the pp32 family of nuclear phosphoproteins, several disease-associated gene products and other characterized and uncharacterized proteins.

Cell Differentiation↗

PubMed2003

Nuclear localization of Galectin-3 in transformed thyroid cells: a role in transcriptional regulation.

The differential proteomic approach (2D gel analysis coupled to MALDI-MS analysis) of nuclear proteins can provide an extremely useful tool to understand control of cell proliferation and differentiation. In order to identify possible markers of dedifferentiation between normal and cancerous thyroid cells, we used a differential proteomics approach by comparing nuclear extracts from the normal rat thyroid cell line FRTL-5 and the completely undifferentiated Ki-mol cell line, obtained by transformation with the Ki-ras oncogene. Galectin-3 (Gal-3) was identified as highly expressed, in the nuclear compartment, only in the transformed cell line. By using different human cancer cell lines, we showed that Gal-3 is maximally expressed in nuclei of papillary cancer cells. We focused on the functional relationship existing between Gal-3 and the thyroid-specific transcription factor TTF-1, whose expression is maintained in papillary cancer where it can contribute to the proliferating status. By using gel-retardation and transient tranfection assays, we demonstrate that Gal-3 upregulates the TTF-1 transcriptional activity. GST-pulldown experiments demonstrate the occurrence of interaction between Gal-3 and TTF-1 homeodomain. Since several lines of evidence suggest a role for Gal-3 in controlling proliferation and tumor progression in thyroid cancer, the stimulatory activity played by Gal-3 over TTF-1 would account for a possible molecular mechanism through which the galectin controls proliferation in thyroid cells.

Animals↗

PubMedDate not supplied

Proteomic analysis of human serum by two-dimensional differential gel electrophoresis after depletion of high-abundant proteins.

Two-dimensional differential gel electrophoresis (2-D DIGE) was used to analyze human serum following the removal of albumin and five other high-abundant serum proteins. After protein removal, serum was analyzed by SDS-PAGE as a preliminary screen, and significant differences between four high-abundant protein removal methods were observed. Antibody-based albumin removal and high-abundant protein removal methods were found to be efficient and specific. To further characterize serum after protein removal, 2-D DIGE was employed, enabling multiplexed analysis of serum through the use of three fluorescent protein dyes. Comparison between crude serum and serum after removal of high-abundant proteins clearly illustrates an increase in the number of lower abundant protein spots observed. Approximately 850 protein spots were detected in crude serum whereas over 1500 protein spots were exposed following removal of six high-abundant proteins, representing a 76% increase in protein spot detection. Several proteins that showed a 2-fold increase in intensity after depletion of high-abundant proteins, as well as proteins that were depleted during abundant protein removal methods, were further characterized by mass spectrometry. This series of experiments demonstrates that high-abundant protein removal, combined with 2-D DIGE, is a practical approach for enriching and characterizing lower abundant proteins in human serum. Consequently, this methodology offers advances in proteomic characterization, and therefore, in the identification of biomarkers from human serum.

Biomarkers↗