The Trypanosoma cruzi Antigen and Epitope Atlas: fine characterization of antibody specificities in human Chagas Disease patients across human populations
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Alejandro Ricci
PROVIDER: E-MTAB-11655 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:These assays represent an antigen discovery screening, and epitope mapping characterization. In this screening two complete proteomes from Trypanosoma cruzi, from two different strains (CL-Brener, Sylvio X10), were displayed in the form of short peptides (tiling array, overlapped) and assayed with pooled serum samples (antibodies) from Chagas Disease patients and matched negative (healthy) subjects selected from 6 geographic regions across the Americas. Peptide arrays (slides) were incubated with pooled serum samples (primary antibodies), washed, and then incubated with a fluorescently-labeled anti-human IgG commercial antibody (secondary antibodies). Raw readouts of fluoresence (signal), as well as normalized signal values are provided in this submission for all samples analyzed. All samples were analyzed in duplicate.
2022-08-20 | E-MTAB-11651 | biostudies-arrayexpress
Project description:Successful vaccination against smallpox with conventional vaccinia virus is usually determined by the development of a vesicular skin lesion at the site of vaccinia inoculation, called a ‘take’. Although previous vaccination is known to be associated with attenuation of the take, the immunology that underlies a no-take in vaccinia-naïve individuals is not well understood. We hypothesized that antibody profiling of individuals before and after receiving vaccinia would reveal differences between takes and no-takes that may help better understand the phenomenon. Using vaccinia proteome microarrays and recombinant protein ELISAs we first examined the antibody response in vaccinia-naïve individuals that failed to take after receiving different doses of the replication competent smallpox vaccines, DryVax® and APSV. Most that received diluted vaccine failed to respond, whereas 4 other no-takes receiving diluted vaccine, and 4 receiving undiluted vaccine, mounted an antibody response. Interestingly, their antibody profiles were not significantly different from controls that did show a take. However, we did find elevated antibody titers in no-takes prior to receiving DryVax® that was significantly different from takes. Although the sample size studied was small, we conclude the failure to take in responders correlates with pre-existing immunity of unknown etiology that may attenuate the skin reaction in a way similar to previous smallpox vaccination. Antibody profiling was peformed on sera from vaccinees receiving smallpox vaccines, either Wyeth DryVax or Aventis Pasteur Vaccine ('WetVax') in NIH-sponsored clinical trials. These samples comprise 23 vaccinia naïve no-takes, and 50 vaccinia naïve takes and 25 previoulsy vaccinated takes as controls. Samples from d0 (day of vaccination) and at the peak of the antibody repsonse (d28) were taken from each donor and probed in triplicate against the proteome arrays.
2012-01-09 | E-GEOD-34931 | biostudies-arrayexpress
Project description:The on-going COVID-19 pandemic requires a deeper understanding of the long-term antibody responses that persist following SARS-CoV-2 infection. To that end, we determined epitope-specific IgG antibody responses in COVID-19 convalescent sera collected at 5 months post-diagnosis and compared that to sera from naïve individuals. Each serum sample was reacted with a high-density peptide microarray representing the complete proteome of SARS-CoV-2 as 15 mer peptides with 11 amino acid overlap and homologs of spike glycoprotein, nucleoprotein, membrane protein, and envelope small membrane protein from related human coronaviruses. Binding signatures were compared between COVID-19 convalescent patients and naïve individuals using the web service tool EPIphany.
2022-02-05 | E-MTAB-10740 | biostudies-arrayexpress
Project description:Leptospirosis is the most widespread zoonotic disease in the world. The lack of an adequate laboratory test is a major barrier for the diagnosis, especially during the early stages of illness, when antibiotic therapy is most effective. Therefore, there is a critical need for an efficient diagnostic test for this life threatening disease. In order to identify new targets that could be used as diagnostic makers for leptopirosis, we constructed a protein microarray chip comprising 61% of Leptospira interrogans proteome and investigated the IgG response against leptospiral antigens from 274 individuals, including 80 acute-, 80 convalescent-phase patients and 114 healthy control subjects from regions with endemic, high endemic and no endemic transmission of leptospirosis. A nitrocellulose line blot assay was performed to validate the accuracy of the protein microarray results. We found 16 antigens that can discriminate between acute cases and healthy individuals from a region with high endemic transmission of leptospirosis, and 18 antigens that distinguish convalescent cases. Some of the antigens identified in this study, such as LipL32, the non-identical domains of the Lig proteins, GroEL and Loa22 are already known to be recognized by sera from human patients, thus serving as a proof-of-concept for the serodiagnostic antigen discovery approach. Several novel antigens were identified, including the hypothetical protein LIC10215 which showed good sensitivity and specificity rates for both acute- and convalescent-phase patients. Our study is the first large-scale evaluation of immunodominant antigens associated with naturally acquired leptospiral infection and novel as well as known serodiagnostic leptospiral antigens that are recognized by antibodies in the sera of leptospirosis cases were identified. The novel antigens identified here may have potential use in both the development of new tests and the improvement of currently available assays for diagnosing this neglected tropical disease. Further research is needed to assess the accuracy of these antigens in more appropriate diagnostic platforms. Antibody profiling was peformed on sera from infected and non-infected subjects to investigate the IgG response against leptospiral antigens. These samples comprised 80 acute-, 80 convalescent-phase patients and 114 healthy control subjects, including 29 subjects from United States (non-endemic area), 35 blood donors from Salvador/Brazil (endemic area) and 50 healthy individuals from region with high endemicity of leptospirosis.
2012-12-04 | E-GEOD-42720 | biostudies-arrayexpress
Project description:Proteins form a diverse array of complexes that mediate cellular function and regulation. A largely unexplored feature of such protein complexes is the selective participation of specific protein isoforms and/or post-translationally modified forms. In this study, we combined native size-exclusion chromatography (SEC) with high-throughput proteomic analysis to characterize soluble protein complexes isolated from human osteosarcoma (U2OS) cells. Using this approach, we have identified over 71,500 peptides and 1,600 phosphosites, corresponding to over 8,000 proteins, distributed across 40 SEC fractions. This represents >50% of the predicted U2OS cell proteome, identified with a mean peptide sequence coverage of 27% per protein. Three biological replicates were performed, allowing statistical evaluation of the data and demonstrating a high degree of reproducibility in the SEC fractionation procedure. Specific proteins were detected interacting with multiple independent complexes, as typified by the separation of distinct complexes for the MRFAP1-MORF4L1-MRGBP interaction network. The data also revealed protein isoforms and post-translational modifications that selectively associated with distinct subsets of protein complexes. Surprisingly, there was clear enrichment for specific Gene Ontology terms associated with differential size classes of protein complexes. This study demonstrates that combined SEC/MS analysis can be used for the system-wide annotation of protein complexes and to predict potential isoform-specific interactions. All of these SEC data on the native separation of protein complexes have been integrated within the Encyclopedia of Proteome Dynamics, an online, multidimensional data-sharing resource available to the community. U2OS cells were grown in DMEM supplemented with 10% FCS, 100 U/l penicillin, and 100 μg/l streptomycin at 37 °C in 10% CO2 and passaged at ~80% confluence. U2OS cells expressing LAP1-tagged MRFAP1 were grown in the same medium but with the addition of 150 μg/ml hygromycin B and 15 μg/ml blasticidine HCl. The cells for each condition were harvested separately via trypsinization, washed in PBS, and lysed in IP buffer (1% Nonidet P-40, 50 mm Tris-HCl, pH 7.4, 10% glycerol, 150 mm NaCl, complete protease inhibitor mixture (Roche Applied Science), PhosStop, 50 mm N-ethylmaleimide). The lysates were sonicated for 10 s at 10% power (three times in total) and then centrifuged for 10 min at 17,000g at 4 °C. Equal protein amounts of each sample were then combined with GFP-trap agarose beads from ChromoTek (Martinsried, Germany) that had been washed once in IP buffer (40 μl of 50% GFP-trap bead slurry per IP) and incubated for 2 h at 4 °C with rotation. The beads were then washed three times with IP buffer via centrifugation at 2,000g for 2 min at 4 °C.
2014-08-14 | PXD001220 | Pride
Project description:Impaired inhibitory signaling underlies the pathophysiology of many neuropsychiatric and neurodevelopmental disorders including autism spectrum disorders and epilepsy. Neuronal inhibition is regulated by synaptic and extrasynaptic γ-aminobutyric acid type A receptors (GABAARs), which mediate phasic and tonic inhibition, respectively. These two GABAAR subtypes differ in their function, ligand sensitivity, and physiological properties. Importantly, they contain different α subunit isoforms: synaptic GABAARs contain the α1-3 subunits whereas extrasynaptic GABAARs contain the α4-6 subunits. While the subunit composition is critical for the distinct roles of synaptic and extrasynaptic GABAAR subtypes in inhibition, the molecular mechanism of the subtype-specific assembly has not been elucidated. To address this issue, we purified endogenous α1- and α4-containing GABAARs from adult murine forebrains and examined their subunit composition and interacting proteins using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) and quantitative analysis. We found that the α1 and α4 subunits form separate populations of GABAARs and interact with distinct sets of binding proteins. We also discovered that the β3 subunit, which co-purifies with both the α1 and α4 subunits, has different levels of phosphorylation on serines 408 and 409 (S408/9) between the two receptor subtypes. To understand the role S408/9 plays in the assembly of α1- and α4-containing GABAARs, we examined the effects of S408/9A (alanine) knock-in mutation on the subunit composition of the two receptor subtypes using LC-MS/MS and quantitative analysis. We discovered that the S408/9A mutation results in the formation of novel α1α4-containing GABAARs. Moreover, in S408/9A mutants, the plasma membrane expression of the α4 subunit is increased whereas its retention in the endoplasmic reticulum is reduced. These findings suggest that S408/9 play a critical role in determining the subtype-specific assembly of GABAARs, and thus the efficacy of neuronal inhibition.
2022-09-23 | PXD036295 | Pride
Project description:Epitope mapping studies aim to identify the binding sites of antibody-antigen interactions to enhance the development of vaccines, diagnostics and immunotherapeutic compounds. However, mapping is a laborious process employing time- and resource-consuming M-bM-^@M-^Xwet benchM-bM-^@M-^Y techniques or epitope prediction software that are still in their infancy. For polymorphic antigens, another challenge is characterizing cross-reactivity between epitopes, teasing out distinctions between broadly cross-reactive responses, limited cross-reactions among variants and the truly type-specific responses. A refined understanding of cross-reactive antibody binding could guide the selection of the most informative subsets of variants for diagnostics and multivalent subunit vaccines. We explored the antibody binding reactivity of sera from human patients and Peromyscus leucopus rodents infected with Borrelia burgdorferi to the polymorphic outer surface protein C (OspC), an attractive candidate antigen for vaccine and improved diagnostics for Lyme disease. We constructed a protein microarray displaying 23 natural variants of OspC and quantified the degree of cross-reactive antibody binding between all pairs of variants, using Pearson correlation calculated on the reactivity values using three independent transforms of the raw data: (1) logarithmic, (2) rank, and (3) binary indicators. We observed that the global amino acid sequence identity between OspC pairs was a poor predictor of cross-reactive antibody binding. Then we asked if specific regions of the protein would better explain the observed cross-reactive binding and performed in silico screening of the linear sequence and 3-dimensional structure of OspC. This analysis pointed to the C-terminal helix of the structure as a major determinant of type-specific cross-reactive antibody binding. We developed bioinformatics methods to systematically analyze the relationship between local sequence/structure variation and cross-reactive antibody binding patterns among variants of a polymorphic antigen, and this method can be applied to other polymorphic antigens for which immune response data is available for multiple variants. Antibody profiling was performed on sera from Borrelia burgdorferi infected and non-infected humans and Peromyscus leucopus rodents against 23 variants of the surface protein OspC . For infected human serum samples, the OspC type of the infecting B. burgdorferi strain is unknown; for experimentally-infected P. leucopus serum samples, it is known. Of human serum samples, 55 were from infected individuals and 25 from naive controls. Of P. leucopus serum samples, 23 were from infected individuals and 7 were from naive controls.
2013-04-12 | E-GEOD-45996 | biostudies-arrayexpress
Project description:We used customized peptide arrays of 163 TXNDC16 peptides to identify immunogenic TXNDC16 epitopes and asked whether discrimination of meningioma and control sera is possible with a specific subset selection of all immunogenic epitopes. CelluSpotsM-bM-^DM-" peptide arrays were manufactured by Intavis (Germany) with 163 TXNDC16 spanning peptides spotted as two replicate subarrays. 15-mer peptides overlapping by 10 amino acids were covalently bound to cellulose membranes with their C-termini. Please note that the non_normalized.txt contains Ch1 Mean values for all samples (each sequence represented in duplicates) and the identifiers in the 'index' column corresponds to those in the 'index' column in raw gpr files; sample data table contains classification values (for each sequence) described in the sample data processing field.
2014-07-01 | E-GEOD-58949 | biostudies-arrayexpress
Project description:Poorly differentiated thyroid carcinomas (PDTC) represent a heterogeneous, aggressive entity, presenting features that suggest a progression from well-differentiated carcinomas. To elucidate the mechanisms underlying such progression and identify novel therapeutical targets, we assessed the genome-wide expression in normal thyroid tissues, well-differentiated thyroid carcinomas and PDTC. RNA were extracted from 2 normal thyroid tissues taken from the opposite lobe of thyroid tumors, and 24 thyroid carcinomas: 5 PDTC, 7 classic papillary thyroid carcinomas (cPTC), 8 follicular variants of PTC (fvPTC) and 4 follicular thyroid carcinomas (FTC). All samples were obtained at time of surgery and immediately frozen in liquid nitrogen. We also hybridized a commercial pool of human thyroid total RNA (BD Bioscience). PTC were screened for BRAF mutations and rearrangements of RET/PTC and, in addition, follicular variants were also analyzed for RAS mutations and PAX8-PPARG rearrangements. FTC were screened for RAS and PAX8-PPARG rearrangements. PDTC were analyzed for BRAF, RAS and PAX8-PPARG genes.
2013-12-10 | E-GEOD-53157 | biostudies-arrayexpress
Project description:ATC are among the most lethal malignancies, for which there is no effective treatment. ATC global gene expression was profiled and compared to normal thyroid tissues, in order to elucidate the molecular alterations contributing to ATC development, and to identify novel therapeutic targets. RNA were extracted from 5 ATC samples. 3 samples were obtained at time of surgery, and were immediately frozen in liquid nitrogen. The RNA from the remaining samples were collected during fine-needle aspiration biopsies, being conserved in RLT buffer (RNeasy Mini Kit, Quiagen) and maintained at -70ºC. RNA were also extracted from 3 normal thyroid samples, which were collected during surgery, taken from the opposite lobe of thyroid tumors. We also hybridized a commercial pool of human thyroid total RNA (BD Bioscience).
2013-12-07 | E-GEOD-53072 | biostudies-arrayexpress