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|
| 1 |
GPL555 |
angiogenesis |
20,800 |
SA Pathology |
2003-10-21 |
|
Homo sapiens
 |
Antibody |
spotted DNA/cDNA, angiogenesis, Cell culture - In vitro capillary tube model: HUVEC were isolated as described previously[Gamble,1993] and cultured in gelatin-coated flasks in M199 medium with Earles balanced salts and 0.68 mM glutamine, 20 mM Hepes, 20% FCS, 15 ug/ml endothelial cell growth supplement (ECGS, BD Biosciences, Bedford, MA, USA), 50 U/ml penicillin, 50 U/ml streptomycin and 15 ug/ml heparin (Sigma, St. Louis, MO, USA). The capillary tube formation assay was performed as previously described[Gamble,1993]. Briefly, 6.4 x 104 cells/160 ul HUVEC medium were plated onto 100 ul gelled bovine type I collagen (Celtrix Laboratories, Palo Alto, CA, USA) in 96-well flat-bottomed microtitre plates. Capillary tube formation was stimulated by the addition of 20 ng/ml phorbol myristate acetate (PMA) and an antibody against A2B1-integrin (RMAC11) which promotes the formation of complex multicellular tubes. Generation of subtracted cDNA libraries: HUVEC from 3 different umbilical cords were plated onto 100ul of bovine type I collagen in 96-well microtitre plates (15 wells/cord). At times 0, 0.5, 3, 6 and 24 hours after stimulation with PMA and RMAC11, total RNA was isolated using the TRIZOL procedure (Invitrogen, Carlsbad, CA, USA). These times were chosen because they approximate time points at which distinct morphological events are known to occur[Gamble,1993;Meyer,1997]. Yields of 7-13ug of total HUVEC RNA were obtained. To achieve enough mRNA for subtraction experiments, the "Switching Mechanism At the 5' end of RNA Transcript" (SMART) PCR cDNA synthesis procedure (Clontech, Palo Alto, CA, USA) was used to convert the mRNA to cDNA and to preferentially amplify full length cDNAs. The PCR Select? cDNA Subtraction Kit (Clontech) was then used to perform suppression subtractive hybridisation between each adjacent time point in both the forward and reverse orientation. Four forward and four reverse libraries were then generated by digesting the subtracted cDNAs with EagI and cloning into the NotI site in pBluescriptIISK+ (Stratagene, La Jolla, CA, USA). These libraries were used to transform XL-10 Gold? ultracompetent bacteria (Stratagene). Generation and probing of microarrays: Generation of microarrays was performed by the Australian Genome Research Facility (AGRF, Melbourne, Australia). Colonies from the forward subtracted libraries (0-0.5h 3200 colonies; 0.5-3h; 3000 colonies, 3-6h, 2800 colonies; 6-24h, 1000 colonies) were robotically picked and glycerols stocks generated. A total of 10,000 clones were picked. Each clone was then PCR amplified using primers to the T7 and T3 promoters in the vector that flank the cloning site. The PCR products were microarrayed onto glass slides in duplicate. The cDNAs used in this microarray were identified as part of collaborative project between the IMVS and Bionomics Limited. The project aims were to identify genes up-regulated during in vitro capillary tube formation as targets for angiogenesis-based therapeutics. Any requests for further information regarding the data generated from this collaboration should be directed to Bionomics Limited (www.bionomics.com.au) at the following address: Bionomics Limited 31 Dalgleish Street Thebarton, South Australia Australia, 5031 Phone: 618 8354 6104 Fax: 618 8354 6199 Email: busdev@bionomics.com.au Keywords = in-vitro Keywords = HUVEC Keywords = capillary tube formation |
| 2 |
GPL173 |
AB-8 |
2,304 |
Stanford Microarray Database (SMD) |
2002-05-22 |
|
Homo sapiens
|
Antibody |
antibody, AB-8, A microarray with 2304 spot features. Tip Configuration: Standard 16-tip Columns per Sector: 12 Rows per Sector: 12 Column Spacing: 175 Row Spacing: 175 |
| 3 |
GPL1909 |
Proteomic analysis of pervanadate-induced tyrosine-phosphorylated proteins in hepatocellular carcinoma WRL 68 cells |
2,110 |
Institute of Biochemistry and Cell Biology |
2005-03-14 |
|
Homo sapiens
|
Antibody |
MS, Proteomic analysis of pervanadate-induced tyrosine-phosphorylated proteins in hepatocellular carcinoma WRL 68 cells, Proteomic analysis of pervanadate-induced tyrosine-phosphorylated proteins in hepatocellular carcinoma WRL 68 cells Protein tyrosine phosphorylation plays critical roles in modulating biological processes such as cellular proliferation, differentiation, migration, apoptosis and metabolism. To profile tyrosine phosphorylated (pTyr) proteins as well as search novel pTyr proteins as cross-talk points among different cellular pathways, we developed a rapid and efficient approach to identify cellular pTyr proteins and their complexes by a combination of subcellular proteomics approach with signal transduction strategies. Human hepatocytic cells from WRL68 cell line were treated with pervanadate (POV), subfractionated into four fractions and then subjected to immunoaffinity purification with anti-pTyr antibody. The eluted mixtures of the anti-pTyr purification were identified by LC-MS/MS. Subcellular fractionation and affinity purification of tyrosine-phosphorylated proteins: WRL68 cells were first grown to 80% confluence in MEM complete medium and then the medium replaced with serum free media. After 15 h, the cells were either untreated or stimulated with 0.1 mM pervanadate (1 mM sodium orthovanadate, 3 mM H2O2) for 10 min. 150-mm cultures of WRL 68 cells were rinsed twice with 4? PBS and then scraped from the dish in 750?l of hypotonic buffer (10 mM Tris, 1 mM NaF, 10 mM IAA, pH 7.5) containing a cocktail of protein inhibitors. After a 20-min incubation on ice, the cells were passed about five times through a 25-g needle. The resulting lysate was subjected to a 15min centrifugation at 1000 rpm at 4?, after which the pellet was resuspended in 250?l of hypotonic buffer and re-extracted by a second round of the trituration and centrifugation. The supernatants of the first and second spins were combined, adjusted to 0.25 M NaCl, and separated into cytosolic (supernatant) and membrane (pellet) fractions bycentrifugation at 19,000 rpm (43,000 g) for 90 min at 4?. All the pellets and total cell lysate were resuspended in RIPA buffer (50 mM Tris-HCl, 150 mM NaCl, 1% Triton X-100, 0.1% SDS, 1% deoxycholic acid sodium) containing 1mM pervanadate with a cocktail of protein inhibitors with sonication aid. Cleared cell lysates were incubated overnight at 4? with 30?l monoclonal anti-phosphotyrosine-agrose (Sigma). Precipitated immune complexes were washed three times with 1×HNTG (20 mM HEPES, 150 mM NaCl, 0.1% Triton X-100, 10% Glycerol, pH 7.5) and then eluted with 100 mM phenyl phosphate (Sigma) in lysis buffer at 4?. Enzyme digestion, mass spectrometry and protein identification: The sample was digested according to the published method. Chromatography was performed using a surveyor LC system (Thermo Finnigan,SanJose,CA) on C18 reverse phase column(RP, 180 µm x 150 mm, BioBasic® C18, 5 µm, Thermo Hypersil-Keystone). The pump flow rate is split 1:100 for a colum flow rate of 1.5 µL/min.The column effluent is directly electrosprayed using the orthogonal metal needle source without further splitting.Mobile phase A is 0.1% formic acid in water,and the B mobile phase is 0.1% formic acid in acetonitrile. The separation of peptides obtained by enzymatic digest of bile sample was achieved with a gradient of 2-80% B over 360 min.The column effluent from the reverse phase column was analyzed by LCQ Deca XP ion-trap mass spectrometer.The micro-electrospray interface uses a 30 µm metal needle that is orthogonal to the inlet of the LCQ.The mass spectrometer was set that one full MS scan was followed by three MS/MS scans on the three most intense ion from the MS spectrum with the following Dynamic Exclusion™ settings: repeat count, 1; repeat duration, 0.5 min; exclusion duration, 3.0 min. The acquired MS/MS spectra were automatically searched against protein database for human proteins (SWISS-PROT/TrEMBL) using the TurboSEQUEST program in the BioWorks™ 3.0 software suite. An accepted SEQUEST result had to have a ?Cn score of at least 0.1 (regardless of charge state) and Xcorr (one charge?1.5, two charges?2.0, three charges?2.5). Single peptides that alone identify a protein were manually validated after meeting the above criteria. Bioinformatics analysis: The pI and MW of the proteins were analyzed using ExPASy proteomics tools accessed from http://cn.expasy.org/tools/#proteome. The grand average hydropathicity (GRAVY) values were determined according to Kyte-Doolittle. The protein subcellular location annotation was from SwissProt &TrEMBL protein database (http://us.expasy.org/sprot/). The protein function family was categorized according to Gene Ontology (GO) annotation terms extracted by InterPro (http://www.ebi.ac.uk./interpro/). The annotation of protein phosphorylation was from SwissProt &TrEMBL protein database (http://us.expasy.org/sprot/) and PhosphoSite (http://www.phosphosite.org). The kinases were annotated according to human kinome. |
| 4 |
GPL172 |
AB-14 |
1,440 |
Stanford Microarray Database (SMD) |
2002-05-22 |
|
Homo sapiens
|
Antibody |
antibody, AB-14, A microarray with 1440 spot features. Tip Configuration: Standard 16-tip Columns per Sector: 10 Rows per Sector: 9 Column Spacing: 375 Row Spacing: 375 |
| 5 |
GPL171 |
AB-11 |
1,152 |
Stanford Microarray Database (SMD) |
2002-05-22 |
|
Homo sapiens
|
Antibody |
antibody, AB-11, A microarray with 1152 spot features. Tip Configuration: Standard 16-tip Columns per Sector: 9 Rows per Sector: 8 Column Spacing: 200 Row Spacing: 200 |
| 6 |
GPL7390 |
AbC ELTE_human_960_v1 |
960 |
ELTE-MTA |
2008-09-26 |
Eotvos Lorand University, Budapest, Hungary |
Homo sapiens
|
Antibody |
antibody, AbC ELTE_human_960_v1, Spotted antigens (protein, peptide, DNA … ) |
| 7 |
GPL16986 |
MUG homo sapiens 8 v KAM 1.2 |
812 |
Medical University Graz |
2013-04-09 |
Kinexus Bioinformatics Corp., Vancouver, Canada, (www.kinexus.ca) |
Homo sapiens
|
Antibody |
antibody, MUG homo sapiens 8 v KAM 1.2, |
| 8 |
GPL13757 |
Kinexus KAM-1.2FN 800 Ab microarray |
800 |
Hospital for Sick Children |
2011-06-21 |
Kinexus Bioinformatics Corportation, Vancouver, BC, Canada |
Homo sapiens
|
Antibody |
antibody, Kinexus KAM-1.2FN 800 Ab microarray, Contains 800 (500 pan-specific and 300 phosphorylation-site specific) antibodies. Most of the antibodies cross-react with mouse. |
| 9 |
GPL120 |
Ab Microarray 380 (Cat. #K1847-1) |
768 |
BD Biosciences Clontech |
2002-04-24 |
|
Homo sapiens
|
Antibody |
antibody, Ab Microarray 380 (Cat. #K1847-1), The BD Clontech Ab Microarray 380 is composed of 378 of distinct, well-characterized monoclonal antibodies covelently immobilized on a glass slide. The antibodies were selected for their specificities and their high binding affinities, and detect a wide variety of intracellular proteins, both cytosolic and membrane-bound, representing a broad range of functional classes. |
| 10 |
GPL9787 |
KINEXUS KAM-1.1 kinome array |
623 |
VU University Medical Center |
2009-12-11 |
KINEXUS BIOINFORMATICS COMPANY, Canada |
Homo sapiens
|
Antibody |
antibody, KINEXUS KAM-1.1 kinome array, phospho-protein-specific antibody array |
| 11 |
GPL10888 |
NYUDM Human 507 antibodies v1.0 |
510 |
New York University |
2010-09-07 |
RayBiotech Inc. |
Homo sapiens
|
Antibody |
antibody, NYUDM Human 507 antibodies v1.0, |
| 12 |
GPL11222 |
mTOR Signaling Phospho Specific Antibody Microarray |
140 |
University of Louisville |
2010-11-17 |
Full Moon Biosystems, Inc. USA |
Homo sapiens
|
Antibody |
antibody, mTOR Signaling Phospho Specific Antibody Microarray, PI3K/mTOR specific Phospho-antibody array |
| 13 |
GPL6245 |
S&S serum biomarker chips, 120 antibodies |
126 |
University of Michigan |
2007-12-05 |
Whatman Schleicher & Schuell |
Homo sapiens
|
Antibody |
antibody, S&S serum biomarker chips, 120 antibodies, |
| 14 |
GPL10337 |
RayBio Homo sapiens 0.1K apoptosis array |
112 |
UMDNJ/NJ Medical School |
2010-04-17 |
RayBiotech, Inc. |
Homo sapiens
|
Antibody |
antibody, RayBio Homo sapiens 0.1K apoptosis array, Principle of the Assay With RayBio® Human Apoptosis Antibody Array kit, researchers can now simultaneously detect the relative level of 43 apoptosis related proteins in cell lysates. By monitoring the changes in protein levels in different experimental model systems, researchers can study pathway activation without spending excess time and effort in performing immunoprecipitations and/or Western Blotting. With RayBio® Human Apoptosis Antibody Array kit, each of the capture antibodies is printed on the membranes, and then treated or untreated cell lysate is added into antibody array membranes. After extensive washing, the membranes are incubated with a cocktail of biotin-conjugated anti-apoptotic protein antibodies. After incubation with HRP-Streptavidin, the signals are visualized by chemiluminescence |
| 15 |
GPL7696 |
Luminex Kinase Phosphorylation Immunosandwich Assay |
69 |
The Broad Institute |
2008-11-24 |
The Broad Institute, Cancer Program, Golub Lab |
Homo sapiens
|
Antibody |
antibody, Luminex Kinase Phosphorylation Immunosandwich Assay, |
| 16 |
GPL14559 |
RayBio Human Cytokine Antibody Array G Series 2000; 6-8 array |
60 |
Medical University Innsbruck |
2011-09-09 |
RayBiotech, Inc. |
Homo sapiens
|
Antibody |
antibody, RayBio Human Cytokine Antibody Array G Series 2000; 6-8 array, Human 6 - 8 arrays.gal |
| 17 |
GPL14560 |
RayBio Human Cytokine Antibody Array G Series 2000; 7-8 array |
60 |
Medical University Innsbruck |
2011-09-09 |
RayBiotech, Inc. |
Homo sapiens
|
Antibody |
antibody, RayBio Human Cytokine Antibody Array G Series 2000; 7-8 array, Human 7 - 8 arrays.gal |
| 18 |
GPL13165 |
Medsaic Human DotScan antibody microarray v3 |
57 |
Brigham and Women's Hospital |
2011-02-14 |
Medsaic; Eveleigh, NSW, Australia |
Homo sapiens
|
Antibody |
spotted peptide or protein, Medsaic Human DotScan antibody microarray v3, |
| 19 |
GPL14561 |
RayBio Human Cytokine Antibody Array G Series 2000; 8-8 array |
54 |
Medical University Innsbruck |
2011-09-09 |
RayBiotech, Inc. |
Homo sapiens
|
Antibody |
antibody, RayBio Human Cytokine Antibody Array G Series 2000; 8-8 array, Human 8 - 8 arrays.gal |
| 20 |
GPL10899 |
NYUDM Human 40 antibodies v1.0 |
42 |
New York University |
2010-09-09 |
RayBiotech Inc. |
Homo sapiens
|
Antibody |
antibody, NYUDM Human 40 antibodies v1.0, Circular feature type |
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