Digital microfluidic hydrogel microreactors for proteomics.

Abstract:

:Proteolytic digestion is an essential step in proteomic sample processing. While this step has traditionally been implemented in homogeneous (solution) format, there is a growing trend to use heterogeneous systems in which the enzyme is immobilized on hydrogels or other solid supports. Here, we introduce the use of immobilized enzymes in hydrogels for proteomic sample processing in digital microfluidic (DMF) systems. In this technique, preformed cylindrical agarose discs bearing immobilized trypsin or pepsin were integrated into DMF devices. A fluorogenic assay was used to optimize the covalent modification procedure for enzymatic digestion efficiency, with maximum efficiency observed at 31 μg trypsin in 2-mm diameter agarose gel discs. Gel discs prepared in this manner were used in an integrated method in which proteomic samples were sequentially reduced, alkylated, and digested, with all sample and reagent handling controlled by DMF droplet operation. Mass spectrometry analysis of the products revealed that digestion using the trypsin gel discs resulted in higher sequence coverage in model analytes relative to conventional homogenous processing. Proof-of-principle was demonstrated for a parallel digestion system in which a single sample was simultaneously digested on multiple gel discs bearing different enzymes. We propose that these methods represent a useful new tool for the growing trend toward miniaturization and automation in proteomic sample processing.

journal_name

Proteomics

journal_title

Proteomics

authors

Luk VN,Fiddes LK,Luk VM,Kumacheva E,Wheeler AR

doi

10.1002/pmic.201100608

subject

Has Abstract

pub_date

2012-05-01 00:00:00

pages

1310-8

issue

9

eissn

1615-9853

issn

1615-9861

journal_volume

12

pub_type

杂志文章
  • SWATH-MS as a tool for biomarker discovery: From basic research to clinical applications.

    abstract::In the era of quantitative proteomics, where mass spectrometry plays a pivotal role, in particular associated with the use of data-independent acquisition, it is time to perform an overview of this growing field with special focus on one of the most promising approaches: SWATH-MS, and to present future perspectives fo...

    journal_title:Proteomics

    pub_type: 杂志文章,评审

    doi:10.1002/pmic.201600278

    authors: Anjo SI,Santa C,Manadas B

    更新日期:2017-02-01 00:00:00

  • Selective and sensitive quantification of the cytochrome P450 3A4 protein in human liver homogenates through multiple reaction monitoring mass spectrometry.

    abstract::This study aimed at establishing a sensitive multiple reaction monitoring-mass spectrometry (MRM-MS) method for the quantification of the drug metabolizing cytochrome P450 (CYP)3A4 enzyme in human liver homogenates. Liver samples were subjected to trypsin digestion. MRM-MS analyses were performed using three transitio...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.201500386

    authors: Cieślak A,Kelly I,Trottier J,Verreault M,Wunsch E,Milkiewicz P,Poirier G,Droit A,Barbier O

    更新日期:2016-11-01 00:00:00

  • A structure-based strategy for discovery of small ligands binding to functionally unknown proteins: combination of in silico screening and surface plasmon resonance measurements.

    abstract::In the postgenomic era, many researchers and organizations have been engaged in structural and functional analyses of proteins. As a part of these efforts, searching for small organic compounds that bind specifically to target proteins is quite important. In this study, we have developed a rational strategy for ligand...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.200401032

    authors: Yoshitani N,Satou K,Saito K,Suzuki S,Hatanaka H,Seki M,Shinozaki K,Hirota H,Yokoyama S

    更新日期:2005-04-01 00:00:00

  • The New Face of the Lipid Droplet: Lipid Droplet Proteins.

    abstract::The lipid droplet (LD) is an organelle with vital functions found in nearly all organisms. LD proteomic research has provided fundamentally important insights into this organelle's functions. The review provides a summary of LD proteomic studies conducted across diverse organisms and cell and tissue types. The accumul...

    journal_title:Proteomics

    pub_type: 杂志文章,评审

    doi:10.1002/pmic.201700223

    authors: Zhang C,Liu P

    更新日期:2019-05-01 00:00:00

  • The up-regulation of 14-3-3 proteins in Smad4 deficient epidermis and hair follicles at catagen.

    abstract::Each postnatal hair follicle (HF) perpetually goes through three phases: anagen, catagen, and telogen. The molecular signals that orchestrate the follicular transition between phases are still largely unknown. Our previous study shows that the keratinocyte specific Smad4 knockout mice exhibit progressive alopecia due ...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.200700760

    authors: Yuan C,Jiao L,Yang L,Ying W,Hu Z,Liu J,Cui F,Li L,Qian L,Teng Y,Hang H,Qian X,Yang X

    更新日期:2008-06-01 00:00:00

  • Native immunoblotting of blue native gels to identify conformation-specific antibodies.

    abstract::A large repertoire of immunological methods permits monitoring the interaction of antibodies with their specific antigen. However, recognition of a protein by a conformation-specific antibody represents a challenge because native conditions must be kept throughout the assay. Native immunoblotting of blue native gels c...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.200900468

    authors: Zickermann V,Wumaier Z,Wrzesniewska B,Hunte C,Schägger H

    更新日期:2010-01-01 00:00:00

  • Mitochondria in metabolic disease: getting clues from proteomic studies.

    abstract::Mitochondria play a key role as major regulators of cellular energy homeostasis, but in the context of mitochondrial dysfunction, mitochondria may generate reactive oxidative species and induce cellular apoptosis. Indeed, altered mitochondrial status has been linked to the pathogenesis of several metabolic disorders a...

    journal_title:Proteomics

    pub_type: 杂志文章,评审

    doi:10.1002/pmic.201300376

    authors: Peinado JR,Diaz-Ruiz A,Frühbeck G,Malagon MM

    更新日期:2014-03-01 00:00:00

  • Translating Cancer Molecular Variability into Personalized Information Using Bulk and Single Cell Approaches.

    abstract::Cancer research is striving toward new frontiers of assigning the correct personalized drug(s) to a given patient. However, extensive tumor heterogeneity poses a major obstacle. Tumors of the same type often respond differently to therapy, due to patient-specific molecular aberrations and/or untargeted tumor subpopula...

    journal_title:Proteomics

    pub_type: 杂志文章,评审

    doi:10.1002/pmic.201900227

    authors: Kravchenko-Balasha N

    更新日期:2020-07-01 00:00:00

  • N-Terminal methylation of proteasome subunit Rpt1 in yeast.

    abstract::The 26S proteasome is a multicatalytic protease complex that degrades ubiquitinated proteins in eukaryotic cells. It consists of a proteolytic core (the 20S proteasome) as well as regulatory particles, which contain six ATPase (Rpt) subunits involved in unfolding and translocation of substrates to the catalytic chambe...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.201300207

    authors: Kimura Y,Kurata Y,Ishikawa A,Okayama A,Kamita M,Hirano H

    更新日期:2013-11-01 00:00:00

  • Comparative proteomics of bacterial pathogens.

    abstract::The monitoring of gene expression via the technologies encompassed under the term 'proteomics' allows proteins of significance to be related to phenotypes associated with strain variability, environmental influences and the effects of genetic manipulation. The characterizations of these molecules are routinely perform...

    journal_title:Proteomics

    pub_type: 杂志文章,评审

    doi:10.1002/1615-9861(200104)1:4<461::AID-PROT461>3.0.

    authors: Cordwell SJ,Nouwens AS,Walsh BJ

    更新日期:2001-04-01 00:00:00

  • Microdissection-An Essential Prerequisite for Spatial Cancer Omics.

    abstract::The problem with cancer tissue is that its intratumoral heterogeneity and its complexity is extremely high as cells possess, depending on their location and function, different mutations, different mRNA expression and the highest intricacy in the protein pattern. Prior to genomic and proteomic analyses, it is therefor...

    journal_title:Proteomics

    pub_type: 杂志文章,评审

    doi:10.1002/pmic.202000077

    authors: von Eggeling F,Hoffmann F

    更新日期:2020-09-01 00:00:00

  • FindPept, a tool to identify unmatched masses in peptide mass fingerprinting protein identification.

    abstract::FindPept (http://www.expasy.org/tools/findpept.html) is a software tool designed to identify the origin of peptide masses obtained by peptide mass fingerprinting which are not matched by existing protein identification tools. It identifies masses resulting from unspecific proteolytic cleavage, missed cleavage, proteas...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/1615-9861(200210)2:10<1435::AID-PROT1435>3

    authors: Gattiker A,Bienvenut WV,Bairoch A,Gasteiger E

    更新日期:2002-10-01 00:00:00

  • Identification of Bacillus anthracis proteins associated with germination and early outgrowth by proteomic profiling of anthrax spores.

    abstract::The use of anthrax spores as a bioweapon has spurred efforts aimed at identifying key proteins expressed in Bacillus anthracis. Because spore germination and outgrowth occur prior to and are required for disease manifestations, blocking germination and early outgrowth with novel vaccines or inhibitors targeting critic...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.200400831

    authors: Huang CM,Foster KW,DeSilva TS,Van Kampen KR,Elmets CA,Tang DC

    更新日期:2004-09-01 00:00:00

  • Global Lysine Crotonylation Profiling of Mouse Liver.

    abstract::Lysine crotonylation (Kcr) is a recently discovered post-translational modification that potentially regulates multiple biological processes. With an objective to expand the available crotonylation datasets, LC-MS/MS is performed using mouse liver samples under normal physiological conditions to obtain in vivo crotony...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.202000049

    authors: Liu JF,Wu SF,Liu S,Sun X,Wang XM,Xu P,Chen HZ,Yang JT

    更新日期:2020-10-01 00:00:00

  • Proteomic profiling of extracellular vesicles secreted from Toxoplasma gondii.

    abstract::Toxoplasma gondii infects a wide range of hosts worldwide, including humans and domesticated animals causing toxoplasmosis disease. Recently, exosomes, small extracellular vesicles (EV) that contain nucleic acids, proteins, and lipids derived from their original cells were linked with disease protection. The effect of...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.201600477

    authors: Wowk PF,Zardo ML,Miot HT,Goldenberg S,Carvalho PC,Mörking PA

    更新日期:2017-08-01 00:00:00

  • Proteomics, nanotechnology and molecular diagnostics.

    abstract::Sequencing of the human genome opened the way to the exploration of the proteome and this has lead to the identification of large numbers of proteins in complex biological samples. The identification of diagnostic patterns in samples taken from patients to aid diagnosis is in the early stages of development. The solut...

    journal_title:Proteomics

    pub_type: 杂志文章,评审

    doi:10.1002/pmic.200700665

    authors: Johnson CJ,Zhukovsky N,Cass AE,Nagy JM

    更新日期:2008-02-01 00:00:00

  • iTRAQ-based quantitative proteomic analysis reveals new metabolic pathways of wheat seedling growth under hydrogen peroxide stress.

    abstract::As an abundant ROS, hydrogen peroxide (H2 O2 ) plays pivotal roles in plant growth and development. In this work, we conducted for the first time an iTRAQ-based quantitative proteomic analysis of wheat seedling growth under different exogenous H2 O2 treatments. The growth of seedlings and roots was significantly restr...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.201300042

    authors: Ge P,Hao P,Cao M,Guo G,Lv D,Subburaj S,Li X,Yan X,Xiao J,Ma W,Yan Y

    更新日期:2013-10-01 00:00:00

  • Membrane-Based SDS Depletion Ahead of Peptide and Protein Analysis by Mass Spectrometry.

    abstract::SDS interferes with both bottom-up and top-down MS analysis, requiring removal prior to detection. Filter-aided sample preparation (FASP) is favored for bottom-up proteomics (BUP) while acetone precipitation is popular for top-down proteomics (TDP). We recently demonstrated acetone precipitation in a membrane filter c...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.201700025

    authors: Unterlander N,Doucette AA

    更新日期:2018-05-01 00:00:00

  • How to distinguish healthy from diseased? Classification strategy for mass spectrometry-based clinical proteomics.

    abstract::SELDI-TOF-MS is rapidly gaining popularity as a screening tool for clinical applications of proteomics. Application of adequate statistical techniques in all the stages from measurement to information is obligatory. One of the statistical methods often used in proteomics is classification: the assignment of subjects t...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.200700046

    authors: Hendriks MM,Smit S,Akkermans WL,Reijmers TH,Eilers PH,Hoefsloot HC,Rubingh CM,de Koster CG,Aerts JM,Smilde AK

    更新日期:2007-10-01 00:00:00

  • A comprehensive dictionary of protein accession codes for complete protein accession identifier alias resolving.

    abstract::In mass spectrometry-based proteomics, protein identification results usually consist of peptide sequences and database-dependent accession identifiers of the matching proteins. Often certain annotations are only available in particular databases that in turn must be queried by a certain identifier. In order to simpli...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.200600018

    authors: Boehm AM,Sickmann A

    更新日期:2006-08-01 00:00:00

  • Proteome analysis of porcine epidemic diarrhea virus (PEDV)-infected Vero cells.

    abstract::Porcine epidemic diarrhea virus (PEDV) causes an acute, highly contagious, and devastating viral enteric disease with a high mortality rate in suckling pigs. A large-scale outbreak of PED occurred in China in 2010, with PEDV emerging in the United States in 2013 and spreading rapidly, posing significant economic and p...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.201400458

    authors: Zeng S,Zhang H,Ding Z,Luo R,An K,Liu L,Bi J,Chen H,Xiao S,Fang L

    更新日期:2015-06-01 00:00:00

  • A glycomic approach to the identification and characterization of glycoprotein function in cells transfected with glycosyltransferase genes.

    abstract::The transfection of glycoprotein glycosyltransferase genes into cells leads to modification of both the structure and function of the glycoproteins and as a result, changes in glycome patterns. N-glycan branching enzymes hold some promise as a model system for the identification of glycome patterns. Both N-acetylgluco...

    journal_title:Proteomics

    pub_type: 杂志文章,评审

    doi:10.1002/1615-9861(200102)1:2<239::AID-PROT239>3.0.

    authors: Taniguchi N,Ekuni A,Ko JH,Miyoshi E,Ikeda Y,Ihara Y,Nishikawa A,Honke K,Takahashi M

    更新日期:2001-02-01 00:00:00

  • Mapping the in vitro interactome of cardiac sodium (Na+ )-calcium (Ca2+ ) exchanger 1 (NCX1).

    abstract::The sodium (Na+ )-calcium (Ca2+ ) exchanger 1 (NCX1) is an antiporter membrane protein encoded by the SLC8A1 gene. In the heart, it maintains cytosolic Ca2+ homeostasis, serving as the primary mechanism for Ca2+ extrusion during relaxation. Dysregulation of NCX1 is observed in end-stage human heart failure. In this st...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.201600417

    authors: Lubelwana Hafver T,Wanichawan P,Manfra O,de Souza GA,Lunde M,Martinsen M,Louch WE,Sejersted OM,Carlson CR

    更新日期:2017-09-01 00:00:00

  • Comparative proteome analysis of porcine follicular fluid and serum reveals that excessive alpha(2)-macroglobulin in serum hampers successful expansion of cumulus-oocyte complexes.

    abstract::Porcine follicular fluid (pFF) constitutes the micro-environment of the maturing oocyte. Although pFF is a transudate of serum, in pigs, it is superior to serum in promoting in vitro expansion of the cumulus cells, a specialized cell population surrounding the oocyte. A comparative proteome analysis of autologous seru...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.200900270

    authors: Bijttebier J,Tilleman K,Dhaenens M,Deforce D,Van Soom A,Maes D

    更新日期:2009-10-01 00:00:00

  • Sarcopenia: Tilting the Balance of Protein Homeostasis.

    abstract::Sarcopenia, defined as age-associated decline of muscle mass and function, is a risk factor for mortality and disability, and comorbid with several chronic diseases such as type II diabetes and cardiovascular diseases. Clinical trials showed that nutritional supplements had positive effects on muscle mass, but not on ...

    journal_title:Proteomics

    pub_type: 杂志文章,评审

    doi:10.1002/pmic.201800411

    authors: Tan KT,Ang SJ,Tsai SY

    更新日期:2020-03-01 00:00:00

  • Range of protein detection by selected/multiple reaction monitoring mass spectrometry in an unfractionated human cell culture lysate.

    abstract::Selected or multiple reaction monitoring is a targeted mass spectrometry method (S/MRM-MS), in which many peptides are simultaneously and consistently analyzed during a single liquid chromatography-mass spectrometry (LC-S/MRM-MS) measurement. These capabilities make S/MRM-MS an attractive method to monitor a consisten...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.201100543

    authors: Ebhardt HA,Sabidó E,Hüttenhain R,Collins B,Aebersold R

    更新日期:2012-04-01 00:00:00

  • Proteomic analysis of the phytopathogenic fungus Botrytis cinerea during cellulose degradation.

    abstract::The ascomycete Botrytis cinerea is a phytopathogenic fungus infecting and causing significant yield losses in a number of crops. Moreover, in the last few years, B. cinerea has been adopted as an important model system in molecular phytopathology. In spite of these contributions, the molecular basis of the infection c...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.200800540

    authors: Fernández-Acero FJ,Colby T,Harzen A,Cantoral JM,Schmidt J

    更新日期:2009-05-01 00:00:00

  • Comparative investigation of seed coats of brown- versus yellow-colored soybean seeds using an integrated proteomics and metabolomics approach.

    abstract::Seed coat color is an important attribute determining consumption of soybean seeds. Soybean cultivar Mallikong (M) has yellow seed coat while its naturally mutated cultivar Mallikong mutant (MM), has brown colored seed coat. We used integrated proteomics and metabolomics approach to investigate the differences between...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.201400453

    authors: Gupta R,Min CW,Kim SW,Wang Y,Agrawal GK,Rakwal R,Kim SG,Lee BW,Ko JM,Baek IY,Bae DW,Kim ST

    更新日期:2015-05-01 00:00:00

  • Comparative proteome and transcriptome analyses of embryonic stem cells during embryoid body-based differentiation.

    abstract::Gene expression analyses of embryonic stem cells (ESCs) will help to uncover or further define signaling pathways and molecular mechanisms involved in the maintenance of self-renewal and pluripotency. We employed a 2-DE-based proteomics approach to analyze human ESC line, Royan H5, in undifferentiated cells and differ...

    journal_title:Proteomics

    pub_type: 杂志文章

    doi:10.1002/pmic.200900003

    authors: Fathi A,Pakzad M,Taei A,Brink TC,Pirhaji L,Ruiz G,Sharif Tabe Bordbar M,Gourabi H,Adjaye J,Baharvand H,Salekdeh GH

    更新日期:2009-11-01 00:00:00

  • Chemoenzymatic method for glycomics: Isolation, identification, and quantitation.

    abstract::Over the past decade, considerable progress has been made with respect to the analytical methods for analysis of glycans from biological sources. Regardless of the specific methods that are used, glycan analysis includes isolation, identification, and quantitation. Derivatization is indispensable to increase their ide...

    journal_title:Proteomics

    pub_type: 杂志文章,评审

    doi:10.1002/pmic.201500266

    authors: Yang S,Rubin A,Eshghi ST,Zhang H

    更新日期:2016-01-01 00:00:00