Project description:To examine changes in tyrosine kinase activity in prostate cancer tumors developing resistance to androgen deprivation therapy and to compare these to changes in tyrosine kinase activity occuring in cell lines exposed to oxygen deficiency (hypoxia)
Project description:In addition to determining possible diagnostic and predictive peptides of lupus and CNS-lupus, we also used our microarray technology along with the Guitope computer program to determine possible natural protein match to five monoclonal autoantibodies that were created using one of the autoimmune MRL/lpr mouse. Submitter states "We have no processed data to submit. We have no gpr files to submit." Microarray analysis was performed on five monoclonal brain-reactive autoantibodies (F9, G10, G4, D1 and D9) that were created from one autoimmune mouse with altered behavior. These samples were tested in duplicated along with a control sample on the GPL17600 platform.
Project description:Immunoglobulin A nephropathy (IgAN) is the most common glomerulonephritis worldwide. However, biomarkers for predicting the progression or regression of IgAN remain a clinical challenge. In the present study, we aim to identify novel diagnostic and prognostic markers of IgAN. Using the cytokine antibody array, we detected serum and urinary levels of nine potential IgAN biomarkers in 32 IgAN patients and 16 healthy controls. The IgAN patients were grouped according to Lee’s grading system, with 16 patients with Grade I–II in the mild IgAN group and 16 with Grade III-V in the severe IgAN group. The associations between levels of these markers and IgAN were evaluated.
Project description:Using this approach the dasatinib-oligonucleotide conjugates was applied to planar arrays of >9,000 human proteins spotted in two technical replicates. All proteins in the commercially available ProtoArray® Human Protein MicroArray (Thermo Fisher Scientific) have been purified and arrayed under native conditions to allow such studies. We adopted this format for investigation of the binding profiles of the drug-oligonucleotide conjugates. Fluorophore-labeled drugs have previously been used to measure binding in protein arrays. The oligonucleotide-conjugated constructed allowed for locally amplified detection via RCA. Circularizing oligonucleotides (padlock probes) were designed with 5’ and 3’ ends complementary to adjacent segments of the oligonucleotides conjugated to the drug molecules. Once converted to oligonucleotide circles by ligation, the probes were replicated through localized RCA, primed by the drug-conjugated oligonucleotides, and visualized using fluorescence-labeled hybridization probes to the repeated sequence of the RCA products. RCA offers a signal enhancement of several hundredfold over singly fluorophore labeled compounds, permitting visualization of even single bound drug probes
Project description:In a two-stage study we investigated levels of Immunoglobulin G (IgG) reactivity in plasma from Myelodysplastic Syndrome (MDS) and Acute Myeloid Leukemia (AML) post MDS patients (59 and 16 cases respectively) as compared to healthy cohort (34). In Stage I we utilized high-throughput protein arrays (23 232 total signals, in duplicate) to identify proteins of high-interest. In Stage II we designed new protein arrays (352 total signals, in duplicate) to further focus on 25 of the proteins from Stage I and expanded to a larger cohort, including both male and female samples (161 MDS and 43 AML patients; 112 healthy controls). Stage I resulted in 35 proteins displaying increased IgG reactivity in patients as compared to the healthy controls (P< 4.3 x10-07, Bonferroni Corrected P<0.01). This protein subset included 14 proteins associated with cancer, 12 with apoptosis, and 3 with the NFAT Regulation canonical pathway. Using the focused arrays we performed a classification of MDS patients and healthy controls. Stage II subsequently identified a high-interest focused set of 3 proteins, namely AKT3, FCGR3A and ARL8B displaying aberrant increased reactivity in patient subgroups, in concordance with Stage I. Autoantibody reactivity against specific proteins provides complementary information to other known molecular signatures for MDS and may enhance our capabilities for detecting and classifying MDS. In the study presented here, MDS patients were classified into Stable, Transforming or AML post MDS (L) classes retrospectively. The different patients were compared to a healthy cohort to assess increased autoantibody reactivity to specific patients as opposed to healthy groups Stage 2 of study: 633 Files analyzed for: 316 Patients in duplicate and a negative control. The proteins on the array were selected following a first stage using ProtoArrays
Project description:A short sequence of 11 amino acids belonging to the cj0669 protein from Campylobacter jejuni NCTC 11168, which was previously identified as potentially immunogenic, was analyzed via alanine scanning to narrow down the significant amino acid residues within the sequence. Twelve peptides, one representing the original sequence and eleven peptides with each residue replaced by alanine in turn, were synthesized on microarrays by JPTs Pepstar Technology. For each microarray, nine replicates for each peptide were spotted. The microarray was separated into three incubation chambers by the ProPlate 3-well module (Grace Biolabs) to allow for incubation with different antibodies in parallel. For specific interactions, rabbit polyclonal IgG to C. jejuni was used, while unspecific binding to the epitope sequence was checked using rabbit polyclonal IgG to Salmonella enterica.
Project description:In search for peptides with higher or special binding affinity and for further understanding of the mode of action, a full substitutional analysis of peptide PeB using microarrays was performed. Thus, 152 PeB mutant variants were generated. In each of them, the full-length sequence was preserved except for only one amino acid from the eight loop-forming amino acids of the original PeB peptide (ARDFYDYDVFYYAMD) which was substituted with the 19 remaining natural amino acids. To assess binding, influenza material was labeled with a protein reacting fluorophore. Microarray-based substitutional analysis of peptide PeB was performed using a PepStar® peptide library spotted on glass slides by JPT Peptide Technologies. The slides were used without additional treatment. For the labeling of proteins with a fluorescent dye, Dyomics DY-634 (λex = 635 nm, λem = 654 nm, Fluoro-spin 634 Kit (emp Biotech) was used, according to the manufacturer´s instructions. The following materials were labeled: NewYork H3N2, Aichi H3N2, Victoria H3N2 and California H1N1. Labeled analytes were incubated several hours or overnight at indicated concentrations using Femtotip buffer (FTP)30 (20 mM Tris, 30% glycerol, 3% polyvinylpyrrolidon 90, 0.1% Tween 20, pH 8.4) for dilution. The slides were washed twice in FTP and twice in ultrapure water and subsequently dried under a stream of nitrogen. Experiments were performed in triplicates using glycans (2,3'-/2,6'-sialyllactose) and proteins (Anti-H1/Anti-H3 antibodies, fetuin) as positive and negative controls.
Project description:This SuperSeries is composed of the following subset Series: GSE33149: Substrate selectivity for semisynthetic CK2 proteins with various posttranslational modifications GSE33150: Substrate selectivity for semisynthetic CK2 proteins with Pin1 Refer to individual Series