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

Antibody isotype epitope mapping of SARS‐CoV‐2 Spike RBD protein: targets for COVID‐19 symptomatology and disease control

Contreras, Marinela; Vicente, Joaquín; Cerón, José Joaquín; Subiela, Silvia Martinez; Urra, José Miguel; Rodríguez‐del‐Río, Francisco J.; Ferreras‐Colino, Elisa; Vaz‐Rodrigues, Rita; de Mera, Isabel G. Fernández; Antunes, Sandra; Domingos, Ana; Gortázar, Christian; de la Fuente, José
Eur J Immunol.
Jan 2023

Humoral Immune Response Profile of COVID-19 Reveals Severity and Variant-Specific Epitopes: Lessons from SARS-CoV-2 Peptide Microarray

Acharjee, Arup; Ray, Arka; Salkar, Akanksha; Bihani, Surbhi; Tuckley, Chaitanya; Shastri, Jayanthi; Agarwal, Sachee; Duttagupta, Siddhartha; Srivastava, Sanjeeva
Viruses.
Jan 2023
The amaranthine scale of the COVID-19 pandemic and unpredictable disease severity is of grave concern. Serological diagnostic aids are an excellent choice for clinicians for rapid and easy prognosis of the disease. To this end, we studied the humoral immune response to SARS-CoV-2 infection to map immunogenic regions in the SARS-CoV-2 proteome at amino acid resolution using a high-density SARS-CoV-2 proteome peptide microarray. The microarray has 4932 overlapping peptides printed in duplicates spanning the entire SARS-CoV-2 proteome. We found 204 and 676 immunogenic peptides against IgA and IgG, corresponding to 137 and 412 IgA and IgG epitopes, respectively. Of these, 6 and 307 epitopes could discriminate between disease severity. The emergence of variants has added to the complexity of the disease. Using the mutation panel available, we could detect 5 and 10 immunogenic peptides against IgA and IgG with mutations belonging to SAR-CoV-2 variants. The study revealed severity-based epitopes that could be presented as potential prognostic serological markers. Further, the mutant epitope immunogenicity could indicate the putative use of these markers for diagnosing variants responsible for the infection.

Antibody Properties Associate with Clinical Phenotype in LGI1 Encephalitis

Ludewig, Susann; Salzburger, Leonie; Goihl, Alexander; Rohne, Jana; Leypoldt, Frank; Bittner, Daniel; Düzel, Emrah; Schraven, Burkhart; Reinhold, Dirk; Korte, Martin; Körtvélyessy, Péter
Cells.
Jan 2023
Autoimmune encephalitis (AE) associated with autoantibodies against leucine-rich glioma-inactivated protein-1 (LGI1) can present with faciobrachial dystonic seizures (FBDS) and/or limbic encephalitis (LE). The reasons for this heterogeneity in phenotypes are unclear. We performed autoantibody (abs) characterization per patient, two patients suffering from LE and two from FBDS, using isolated antibodies specified with single amino acid epitope mapping. Electrophysiological slice recordings were conducted alongside spine density measurements, postsynaptic Alpha-amino-3-hydoxy-5-methyl-4-isoaxole-proprionate-receptors (AMPA-R) and N-methyl-D-aspartate-receptors receptor (NMDA-R) cluster counting. These results were correlated with the symptoms of each patient. While LGI1 abs from LE patients mainly interacted with the Leucine-rich repeat section of LGI1, abs from both FBDS patients also recognized the Epitempin section as well. Six-hour incubation of mouse hippocampal slices with LE patients autoantibodies but not from the FBDS patients resulted in a significant decline in long-term potentiation (p = 0.0015) or short-term plasticity at CA3-CA1 neurons and in decreased hippocampal synaptic density. Cluster differentiation showed no decrease in postsynaptic AMPA-R and NMDA-R. LGI1 autoantibodies selected by phenotype show an almost distinct epitope pattern, elicit disparate functional effects on hippocampal neurons, and cause divergent effects on spine density. This data illuminates potential pathomechanisms for disease heterogeneity in LGI1 AE.

Immunodominant antibody responses directed to SARS-CoV-2 hotspot mutation sites and risk of immune escape

Oliveira, Jamille Ramos; Ruiz, Cesar Manuel Remuzgo; Machado, Rafael Rahal Guaragna; Magawa, Jhosiene Yukari; Daher, Isabela Pazotti; Urbanski, Alysson Henrique; Schmitz, Gabriela Justamante Händel; Arcuri, Helen Andrade; Ferreira, Marcelo Alves; Sasahara, Greyce Luri; de Medeiros, Giuliana Xavier; Júnior, Roberto Carlos Vieira Silva; Durigon, Edison Luiz; Boscardin, Silvia Beatriz; Rosa, Daniela Santoro; Schechtman, Deborah; Nakaya, Helder I.; Cunha-Neto, Edecio; Gadermaier, Gabriele; Kalil, Jorge; Coelho, Verônica; Santos, Keity Souza
Front. Immunol..
Jan 2023
Introduction Considering the likely need for the development of novel effective vaccines adapted to emerging relevant CoV-2 variants, the increasing knowledge of epitope recognition profile among convalescents and afterwards vaccinated with identification of immunodominant regions may provide important information. Methods We used an RBD peptide microarray to identify IgG and IgA binding regions in serum of 71 COVID-19 convalescents and 18 vaccinated individuals. Results We found a set of immunodominant RBD antibody epitopes, each recognized by more than 30% of the tested cohort, that differ among the two different groups and are within conserved regions among betacoronavirus. Of those, only one peptide, P44 (S415-429), recognized by 68% of convalescents, presented IgG and IgA antibody reactivity that positively correlated with nAb titers, suggesting that this is a relevant RBD region and a potential target of IgG/IgA neutralizing activity. Discussion This peptide is localized within the area of contact with ACE-2 and harbors the mutation hotspot site K417 present in gamma (K417T), beta (K417N), and omicron (K417N) variants of concern. The epitope profile of vaccinated individuals differed from convalescents, with a more diverse repertoire of immunodominant peptides, recognized by more than 30% of the cohort. Noteworthy, immunodominant regions of recognition by vaccinated coincide with mutation sites at Omicron BA.1, an important variant emerging after massive vaccination. Together, our data show that immune pressure induced by dominant antibody responses may favor hotspot mutation sites and the selection of variants capable of evading humoral response.

Deciphering the Autoantibody Response to the OJ Antigenic Complex

Fritzler, Marvin J.; Bentow, Chelsea; Satoh, Minoru; McHugh, Neil; Ghirardello, Anna; Mahler, Michael
Diagnostics.
Jan 2023
(1) Background: Myositis specific antibodies (MSA) are important diagnostic biomarkers. Among the rarest and most challenging MSA are anti-OJ antibodies which are associated with anti-synthetase syndrome (ASS). In contrast to the other tRNA synthetases that are targets of ASS autoantibodies (e.g Jo-1, PL-7, PL-12, EJ, KS, Zo), OJ represents a macromolecular complex with several ribonucleoprotein subunits. Therefore, the choice of the antigen in autoantibody assays can be challenging. (2) Methods: We collected two independent cohorts with anti-OJ antibodies, one based on a commercial line immunoassay (LIA) (n = 39), the second based on protein immunoprecipitation (IP) (n = 15). Samples were tested using a particle-based multi-analyte technology (PMAT) system that allows for the simultaneous detection of antibodies to various autoantigens. For the detection of anti-OJ antibodies, two different antigens were deployed (KARS, IARS) on PMAT. The reactivity to the two antigens KARS and IARS was analyzed individually and combined in a score (sum of the median fluorescence intensities). (3) Results: In the cohort selection based on LIA, 3/39 (7.7%) samples were positive for anti-KARS and 7/39 (17.9%) for anti-IARS and 14/39 (35.9%) when the two antigens were combined. In contrast, in samples selected by IP the sensitivity of anti-KARS was higher: 6/15 (40.0%) samples were positive for anti-KARS, 4/15 (26.7%) for anti-IARS and 12/15 (80.0%) for the combination of the two antigens. 18/39 (46.2%) of the LIA samples generated a cytoplasmic IIF pattern (compatible with anti-synthetase antibodies), but there was no association with the antibody levels, neither with LIA nor with PMAT. (4) Conclusions: The combination of IARS and KARS might represent a promising approach for the detection of anti-OJ antibodies on a fully automated platform.

Compounds and Methods Targeting Interleukin-19

Higgs Jr., Richard Earl; Konrad, Robert John; Nickoloff, Brian Jeffrey; Siegel II, Robert William; Mertz, Prema Maria
Nov 2020
The present invention provides compounds and methods targeting human interleukin-19, including therapeutic antibodies, pharmaceutical compositions and diagnostic applications useful in the field of immune-mediated diseases including psoriasis, atopic dermatitis, psoriatic arthritis, bronchial asthma and diabetic nephropathy.

SARS-CoV-2 epitope mapping on microarrays highlights strong immune-response to N protein region

Musicò, Angelo; Frigerio, Roberto; Mussida, Alessandro; Barzon, Luisa; Sinigaglia, Alessandro; Riccetti, Silvia; Gobbi, Federico; Piubelli, Chiara; Bergamaschi, Greta; Chiari, Marcella; Gori, Alessandro; Cretich, Marina

Abstract

A workflow for SARS-CoV-2 epitope discovery on peptide microarrays is herein reported. The process started with a proteome-wide screening of immunoreactivity based on the use of a high-density microarray followed by a refinement and validation phase on a restricted panel of probes using microarrays with tailored peptide immobilization through a click-based strategy. Progressively larger, independent cohorts of Covid-19 positive sera were tested in the refinement processes, leading to the identification of immunodominant regions on SARS-CoV-2 Spike (S), Nucleocapsid (N) protein and Orf1ab polyprotein. A summary study testing 50 serum samples highlighted an epitope of the N protein (region 155-171) providing 92% sensitivity and 100% specificity of IgG detection in Covid-19 samples thus being a promising candidate for rapid implementation in serological tests.

Generation of Chicken IgY against SARS-COV-2 Spike Protein and Epitope Mapping

Lu, Yan; Wang, Yajun; Zhang, Zhen; Huang, Jingliang; Yao, Meicun; Huang, Guobin; Ge, Yuanyuan; Zhang, Peichun; Huang, Huaxin; Wang, Yong; Li, Huiliang; Wang, Wen
Oct 2020
This new decade has started with a global pandemic of COVID-19 caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), precipitating a worldwide health crisis and economic downturn. Scientists and clinicians have been racing against time to find therapies for COVID-19. Repurposing approved drugs, developing vaccines and employing passive immunization are three major therapeutic approaches to fighting COVID-19. Chicken immunoglobulin Y (IgY) has the potential to be used as neutralizing antibody against respiratory infections, and its advantages include high avidity, low risk of adverse immune responses, and easy local delivery by intranasal administration. In this study, we raised antibody against the spike (S) protein of SARS-CoV-2 in chickens and extracted IgY (called IgY-S) from egg yolk. IgY-S exhibited high immunoreactivity against SARS-CoV-2 S, and by epitope mapping, we found five linear epitopes of IgY-S in SARS-CoV-2 S, two of which are cross-reactive with SARS-CoV S. Notably, epitope SIIAYTMSL, one of the identified epitopes, partially overlaps the S1/S2 cleavage region in SARS-CoV-2 S and is located on the surface of S trimer in 3D structure, close to the S1/S2 cleavage site. Thus, antibody binding at this location could physically block the access of proteolytic enzymes to S1/S2 cleavage site and thereby impede S1/S2 proteolytic cleavage, which is crucial to subsequent virus-cell membrane fusion and viral cell entry. Therefore, the feasibility of using IgY-S or epitope SIIAYTMS-specific IgY as neutralizing antibody for preventing or treating SARS-CoV-2 infection is worth exploring.

Ns2b as Marker for Zika Virus Infections

Jaenisch, Thomas; Fischer, Nico; Loeffler, Felix; Sekul, Renate; Stadler, Volker; Marques, Ernesto T. A.; Lucas, Zachariah
Oct 2020
The present invention relates to protein NS2b or fragment(s) thereof as biomarker or diagnostic marker for the diagnosis and/or prognosis of Zika virus infections. The present invention further relates to peptides and cyclic peptides, compositions and arrays and multimer compounds comprising them. The present invention further relates to a method for the diagnosis and/or prognosis of Zika virus infections, comprising the use of protein NS2b or fragment(s) thereof, or of the peptides, cyclic peptides, compositions and/or arrays in immunoassays. The present invention further relates to peptide-based compounds comprising at least one fragment of protein NS2b and at least one further component and to methods for the diagnosis and/or prognosis of Zika virus infections.

Monoclonal Antibodies to Growth and Differentiation Factor 15 (gdf-15), and Uses Thereof for Treating Cancer Cachexia and Cancer

Wischhusen, Jörg; Junker, Markus; Schäfer, Tina; Pühringer, Dirk; Lucas, Zachariah
Oct 2020
The present invention relates to monoclonal anti-human-GDF-15 antibodies. The antibodies include chimeric anti-bodies and humanized antibodies. The invention also relates to monoclonal anti-human-GDF-15 antibodies including murine anti-bodies, chimeric antibodies and humanized antibodies for use in methods for the treatment of cancer cachexia and also for the treatment of cancer. The invention also provides pharmaceutical compositions, kits, methods and uses and cell lines capable of producing the monoclonal antibodies of the invention.

Novel Anti-Cd40 Antibodies and Use Thereof

Park, Chung Gyu; Kim, Jung Sik; Lucas, Zachariah
Sep 2020
The present invention relates to novel anti-CD40 antibodies and a use thereof and, more specifically, provided are a pharmaceutical composition for treating or preventing autoimmune diseases and a composition for inhibiting immune rejection during organ transplantation, both compositions containing, as an active ingredient, novel anti-CD40 antibodies that specifically bind to a novel epitope of CD40. Novel anti-CD40 antibodies of the present invention directly target CD40, but not CD40 ligands, and block the signaling of CD40-CD154 without stimulating platelets so as to exhibit excellent antagonistic effects, thereby being expected to be usable as a preparation effective in the treatment of autoimmune diseases and the inhibition of organ transplantation rejection.

Detection of specific IgE against linear epitopes from Gal d 1 has additional value in diagnosing hen’s egg allergy in adults

Ehlers, Anna M.; Otten, Henny G.; Wierzba, Eva; Flügge, Ulrike; Le, Thuy-My; Knulst, André C.; Suer, Waltraud
Background Although hen’s egg allergy is more prevalent in children, up to 0.6% of adults from different European countries suffers from a persistent or newly-onset hen’s egg allergy, making accurate diagnosis in adults necessary. However, sensitisation to hen’s egg extracts, components and linear epitopes are solely studied in children. Methods Hen’s egg allergic (n=16) and tolerant (n=20) adults were selected by sensitisation towards recombinant components rGal d 1 and/or 3. Sensitisation profiles towards egg white and yolk extract and the native components Gal d 1, 2, 3 and 4 were respectively evaluated with the ImmunoCAP or the EUROLINE system. Characterisation of linear epitopes was performed with a peptide microarray containing 15mer peptides representing the entire sequence of mature Gal d 1 and 3. Results Overall, sIgE titres against hen’s egg extracts and single components overlapped largely between allergic and tolerant adults. Although the median sIgE/sIgG4 ratio to Gal d 1 was increased in allergic adults, the range was comparable between both groups. Clinically relevant sensitisation to Gal d 1 was confirmed by sIgE-binding to the linear epitopes aa30-41, aa39-50 or aa84-95 in 6/13 allergic adults, mainly suffering from objective symptoms. In comparison, these epitopes were recognized by 1/15 tolerant patient. Only a few linear epitopes were detected for Gal d 3, suggesting a greater importance of conformational epitopes for the recognition of Gal d 3. Conclusion and Clinical Relevance Specific IgE-binding to linear epitopes of Gal d 1 is highly specific in identifying hen’s egg allergic adults with objective symptoms.

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