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Discover how PEPperPRINT Peptide Microarray products have been used in different fields of research.

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Protein microarrays for COVID-19 research: Biomarker discovery, humoral response, and vaccine targets

Acharjee, Arup; Barpanda, Abhilash; Ren, Jing; Yu, Xiaobo
Jan 2022
10.1201/9781003220787-6
Of all the technological interventions used to probe the COVID-19 biological sample, microarrays have provided unique information about the biology of SARS-CoV-2 infection in the greatest of detail. Protein microarrays are available in various formats such as protein microarray, antibody microarray, and peptide microarrays. These provide an attractive format to study host response against infection, with its straightforward sample preparation strategy and easy result analysis pipelines. Microarray technology either uses antibodies against hundreds of proteins to study host proteins or scans immunogenic peptides of the pathogen in a microarray panel of the pathogen proteome. It can be used to study the humoral immune response against antigenic proteins of the SARS-CoV-2 virus, host proteomic alterations due to the infection. The SARS-CoV-2 peptide array can be used for the accurate detection of antigenic determinants for vaccine design. This chapter summarizes the different types of protein and peptide microarray and their use in COVID-19 biomarker discovery, disease management, vaccine design, etc., for better management of COVID-19.

Distinct early IgA profile may determine severity of COVID-19 symptoms: an immunological case series

Dahlke, Christine; Heidepriem, Jasmin; Kobbe, Robin; Santer, Rene; Koch, Till; Fathi, Anahita; Ly, My L.; Schmiedel, Stefan; Seeberger, Peter H.; ID-UKE COVID-19 study group; Addo, Marylyn M.; Loeffler, Felix F.
Apr 2020
10.1101/2020.04.14.20059733
SARS-CoV-2 is the causative agent of COVID-19 and is a severe threat to global health. Patients infected with SARS-CoV-2 show a wide range of symptoms and disease severity, while limited data is available on its immunogenicity. Here, the kinetics of the development of SARS-CoV-2-specific antibody responses in relation to clinical features and dynamics of specific B-cell populations are reported. Immunophenotyping of B cells was performed by flow cytometry with longitudinally collected PBMCs. In parallel, serum samples were analyzed for the presence of SARS-CoV-2-specific IgA, IgG, and IgM antibodies using whole proteome peptide microarrays. Soon after disease onset in a mild case, we observed an increased frequency of plasmablasts concomitantly with a strong SARS-CoV-2-specific IgA response. In contrast, a case with more severe progression showed a delayed, but eventually very strong and broad SARS-CoV-2-specific IgA response. This case study shows that determining SARS-CoV-2-specific antibody epitopes can be valuable to monitor the specificity and magnitude of the early B-cell response, which could guide the development of vaccine candidates. Follow-up studies are required to evaluate whether the kinetics and strength of the SARS-CoV-2-specific IgA response could be potential prognostic markers of viral control.

Genomics-Driven Immunoproteomics: An Integrative Platform to Uncover Important Biomarkers for Human Diseases

Giri, Raghavendra; Qendro, Veneta; Rani, Pooja; Jepchumba, Carren; Bugos, Grace; Stadler, Volker; Han, David K.
Jul 2019
10.1007/978-1-4939-9597-4_21
Genomics-driven immunoproteomics (GDI) is a platform that helps identify antigenic protein targets of mutations and other deoxyribonucleic acid (DNA) variations that are commonly associated with pathological states. This platform utilizes data generated from deep sequencing of exomic DNA or ribonucleic acid (RNA) as input to synthesize mutant peptides into microarrays, which then can be used to detect antigenic proteins that invoke immune response in patients. The technology has been used to detect antigenic targets of multiple sclerosis, an autoimmune disease [1], and cancer to identify mutant proteins that invoke immune response in breast cancer patients [2]. This technology has many potential applications to select genomic changes that are specifically recognized by the immune system in a rapid and efficient manner.

192 Disparities of B-cell type I interferon production and responses in SLE

Winn Chatham, W; Hsu, Hui-Chen; Mountz, John; Wu, Qi; Essman, Alex; Ojo, Oluwagbemiga; Liu, Shanrun; Yang, PingAr; Luo, Bao; Hamilton, Jennie
Apr 2019
10.1136/lupus-2019-lsm.192
Background Dysregulated responses to type I interferons (IFNs) is a hallmark of autoreactive B-cell development in SLE. This study investigated the source of IFN, the major type I IFN responsive B cells, and the disparities associated with B-cell IFN production and type I IFN responses. Methods IFN expression in B, CD4 T and plasmacytoid dendritic cells (pDCs) in PBMCs were analyzed by flow cytometry. Single cell gene expression analysis was carried out using the Fluidigm/BioMark system for targeted expression of low abundance genes, and the 10x Chromium platform for unbiased transcriptome and BCR V(D)J analysis of approximately 2,000 B cells per subject. Autoantigen epitope targets were analyzed using a 4287 high-throughput PEPperPrint Autoimmune Epitope Microarray and a conventional ELISA analysis. Results IFN was analyzed in B cells, CD4 T cells and pDCs in PBMCs of SLE patients and healthy controls (HCs). Endogenous IFN was significantly increased in transitional (Tr), mature naïve, and memory B cells of SLE patients compared to HCs. Endogenous IFN in B cells was equivalent to that in pDCs. B-cell endogenous IFN was highly correlated with renal disease, anti-dsDNA, anti-Sm and anti-SSA. Strikingly, the highest correlation of IFN with clinical manifestations was observed in African-American (AA) patients with IgG autoAbs against snRNP323-339, U1snRNP-C97-113. At the single cell transcriptome levels, Tr B cells could be divided into type I IFN expressing (IFN+) or type I IFN stimulated gene (ISG+) subpopulations. TLR7 and TLR3 were mainly expressed by IFN +cells whereas TLR9 was mainly expressed by ISG +B cells. Unbiased single cells analysis of B cells indicated highly expressed ISG gene set in IGHM+, IGHD+, and IGHG +B cells in AA patients with autoantibodies and renal disease. Further, ISG highly expressing SLE B cells exhibited unique heavy- and light-chain repertoires including expression of the autoreactive IGHV4-34 gene, targeted with the 9 G4 anti-idiotype antibody that recognizes DNA- and RBP-autoreactive B cells. Conclusions (i) B cells are an important source of type I IFNs in modulating TLR and BCR responses in SLE; (ii) there are well-orchestrated distinct programs in type I IFN expression and response genes in subsets of B cells, (iii) distinct pathways of autoreactive B cell survival and activation are effected by combined signaling through BCR, TLR, and IFNAR with resultant distinct BCR heavy- and light-chain repertoire.

Automated laser-assisted synthesis of microarrays for infectious disease research

Paris, Grigori; Heidepriem, Jasmin; Tsouka, Alexandra; Mende, Marco; Eickelmann, Stephan; Loeffler, Felix F.
Mar 2019
10.1117/12.2516781
We developed a next-generation method for chemical in–situ combinatorial biomolecule array synthesis. This allows for an unprecedented combinatorial freedom in the automated chemical synthesis of molecule arrays with very high spot densities. Key feature of this new method is an automated positioning and laser transfer process: Small solid material spots are rapidly transferred from a donor film to an acceptor surface, requiring only minute amounts of materials. The transfer is performed with different and easy-to-produce donor slides. Each donor slide bears a thin polymer film, embedding one type of monomer. The coupling reaction occurs in a separate heating step, where the matrix becomes viscous and building blocks can diffuse within the material and couple to the acceptor surface. Since these transferred material spots are only several nanometers thin, this method allows for a consecutive multi-layer material deposition of e.g. activation reagents and amino acids. Subsequent heat-induced mixing facilitates an in–situ activation and coupling of the monomers. Positioning several of such resin spots, containing different chemical reagents, on top of each other, will enable for the first time in such small dimensions unique chemical synthesis strategies for each spot. Amount and concentration of the deposited materials can be adjusted with the laser parameters. Employing similar arrays, we can analyze the human immune response towards the proteome of different pathogens. We screened several peptide array replicas with different patient sera. The screenings resulted in significant hits in several proteins with interesting implications for future diagnostics and vaccine development.

High-Density Peptide Arrays for Malaria Vaccine Development

Loeffler, Felix F.; Pfeil, Johannes; Heiss, Kirsten
Apr 2016
10.1007/978-1-4939-3387-7_32
The development of an efficacious and practicable vaccine conferring sterile immunity towards a Plasmodium infection represents a not yet achieved goal. A crucial factor for the impact of a given anti-plasmodial subunit vaccine is the identification of the most potent parasitic components required to induce protection from both infection and disease. Here, we present a method based on a novel high-density peptide array technology that allows for a flexible readout of malaria antibodies. Peptide arrays applied as a screening method can be used to identify novel immunogenic antibody epitopes under a large number of potential antigens/peptides. Ultimately, discovered antigen candidates and/or epitope sequences can be translated into vaccine prototype design. The technology can be further utilized to unravel antibody-mediated immune responses (e.g., involved in the establishment of semi-immunity) and moreover to confirm vaccine potency during the process of clinical development by verifying the induced antibody responses following vaccination.

A Novel Combinatorial Approach to High-Density Peptide Arrays

Beyer, Mario; Block, Ines; König, Kai; Nesterov, Alexander; Fernandez, Simon; Felgenhauer, Thomas; Schirwitz, Christopher; Leibe, Klaus; Bischoff, Ralf F.; Breitling, Frank; Stadler, Volker
Jan 2009
10.1007/978-1-60327-394-7_16
Combinatorial synthesis of peptides on solid supports (1), either as spots on cellulose membranes (2) or with split-pool-libraries on polymer beads (3), substantially forwarded research in the field of peptide-protein interactions. Admittedly, these concepts have specific limitations, on one hand the number of synthesizable peptide sequences per area, on the other hand elaborate decoding/encoding strategies, false-positive results and sequence limitations. We recently established a method to produce high-density peptide arrays on microelectronic chips (4). Solid amino acid microparticles were charged by friction and transferred to defined pixel electrodes onto the chip’s surface, where they couple to a functional polymer coating simply upon melting (Fig. 16.1 A-D,F). By applying standard Fmoc chemistry according to Merrifield, peptide array densities of up to 40,000 spots per square centimetre were achieved (Fig. 16.1G). The term Merrifield synthesis describes the consecutive linear coupling and deprotecting of L-amino acids modified with base-labile fluorenylmethoxy (Fmoc) groups at the N-terminus and different acid-sensitive protecting groups at their side chains. Removing side chain protecting groups takes place only once at the very end of each synthesis and generates the natural peptide sequence thereby.

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