Contact
Quote

Home » Publications

Publications

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

Filter by
Filtered (38)
Filters
Show (38)
Cancel
Showing most recent

Severity-dependent IgG epitope profiling in COVID-19 reveals differential recognition of pathogen-derived antigens

Do Nascimento, Lais Alves; Machado, NicolleRakanidis; Borges, João Vitor Da Silva; Fagundes, Beatriz Oliveira; Bergamasco, Isabella Siuffi; Sgnotto, Fabio Da Ressureição; Bachi, André Luis Lacerda; Sato, Maria Notomi; Victor, Jefferson Russo
Front. Immunol..
Sep 2025
10.3389/fimmu.2025.1668223
Background The contribution of antibody-mediated responses to COVID – 19 outcomes remains unclear, particularly regarding cross-reactivity with unrelated pathogens. While co-infections are known to influence disease progression, the broader landscape of IgG reactivity during SARS-CoV-2 infection has not been systematically explored. Methods We employed a high-density peptide microarray containing 4,344 linear epitopes from 37 viruses, 27 bacteria, 17 parasites, and 8 fungi to characterize serum IgG repertoires from individuals with moderate (n = 39) or severe (n = 40) COVID – 19. Controls included pre-pandemic healthy donors and a pooled intravenous immunoglobulin (IVIg) formulation. Data analysis included intensity ranking, epitope mapping, and comparative analysis of mean signal intensities for each epitope between the COVID-Mod and COVID-Sev groups. Results COVID – 19 patients showed widespread IgG reactivity against diverse pathogens, with patterns differing by disease severity. Severe cases displayed broader and more intense reactivity, notably against hepatitis C virus (HCV), SARS-CoV-1, influenza A, Mycobacterium tuberculosis, and Plasmodium falciparum. Moderate cases showed preferential recognition of epitopes from HTLV-I, Neisseria meningitidis, and Trypanosoma cruzi. These findings suggest that SARS-CoV-2 infection modulates pre-existing humoral memory, possibly through epitope spreading or immune reprogramming. Conclusions SARS-CoV-2 infection reshapes the IgG epitope repertoire in a severity-dependent manner, extending to antigens from unrelated pathogens. This phenomenon may reflect underlying immune dysregulation or idiotype-driven interactions. Comprehensive profiling of pathogen-related IgG responses may reveal potential biomarkers of disease severity. This phenomenon may inform future investigations aimed at improving personalized management strategies for co-infected or immunocompromised patients.

A tumor-binding antibody with cross-reactivity to viral antigens

Campa, Michael J.; Gottlin, Elizabeth B.; Wiehe, Kevin; Patz, Edward F.
Cancer Immunol Immunother.
Feb 2025
10.1007/s00262-025-03975-8
**Background** We previously identified in non-small cell lung cancer (NSCLC) patients an autoantibody to complement factor H (CFH) that is associated with non-metastatic disease and longer time to progression in patients with stage I disease. A recombinant human antibody, GT103, was cloned from single B cells isolated from patients with the autoantibody. GT103 inhibits tumor growth and establishes an antitumor microenvironment. The anti-CFH autoantibody and GT103 recognize the epitope PIDNGDIT within the SCR19 domain of CFH. Here, we asked if this autoantibody could have originally arisen as a humoral response to a similar epitope in a viral protein from a prior infection. **Methods** Homologous viral peptides with high sequence identity to the core PIDNGDIT epitope sequence were identified and synthesized. NSCLC patient plasma containing anti-CFH autoantibodies were assayed by ELISA against these peptides. GT103 was assayed on a 4345-peptide pathogen microarray. **Results** Epitopes similar to the GT103 epitope are present in several viruses, including human metapneumovirus-1 (HMPV-1) that contains a sequence within attachment glycoprotein G that differs by one amino acid. Anti-CFH autoantibodies in NSCLC patient plasma weakly bound to an HMPV-1 peptide containing the epitope. GT103 cross-reacted with multiple viral epitopes on a peptide microarray, with the top hits being peptides in the human endogenous retrovirus-K polymerase (HERV-K pol) protein and measles hemagglutinin glycoprotein. GT103 bound the viral HMPV-1, HERV-K pol, and measles epitope peptides but with lower affinity compared to the GT103 epitope peptide. **Conclusion** These findings suggest that memory B cells against a viral target could have affinity matured to produce an antibody that recognizes a similar epitope on tumor cells and exhibits antitumor properties.

Investigation of Immunoreactivity Profiles and Epitope Landscape in Divergent COVID-19 Trajectories and SARS-CoV-2 Variants

Bihani, Surbhi; Ray, Arka; Borishetty, Dhanush; Tuckley, Chaitanya; Salkar, Akanksha; Acharjee, Arup; Shrivastav, Prithviraj; Shrivastav, Om; Shastri, Jayanthi; Agrawal, Sachee; Duttagupta, Siddhartha; Srivastava, Sanjeeva
J. Proteome Res..
Jan 2025
10.1021/acs.jproteome.4c00791
This study aimed to elucidate the complexity of the humoral immune response in COVID-19 patients with varying disease trajectories using a SARS-CoV-2 whole proteome peptide microarray chip. The microarray, containing 5347 peptides spanning the entire SARS-CoV-2 proteome and key variants of concern, was used to analyze IgG responses in 10 severe-to-recovered, 9 nonsevere-to-severe cases, and 10 control case (5 pre-pandemic and 5 SARS-CoV-2-negative) plasma samples. We identified 1151 IgG-reactive peptides corresponding to 647 epitopes, with 207 peptides being cross-reactive across 124 epitopes. Nonstructural protein 3 (nsp3) exhibited the highest number of total and unique epitopes, followed by the spike protein. nsp12 had the most number of cross-reactive epitopes. Peptides from the spike protein and nsps 2, 3, 5, and 13 were notably associated with recovery. Additionally, specific mutations in SARS-CoV-2 variants were found to alter peptide immunoreactivity, with some mutations (e.g., G142D, L452R, and N501Y) enhancing and others (e.g., R190S and E484 K) reducing immune recognition. These findings have critical implications for the development of diagnostics, vaccines, and therapeutics. Understanding the distribution of epitopes and the impact of viral mutations on antigenicity provides insights into immune evasion mechanisms, informing strategies for controlling COVID-19 and future coronavirus outbreaks.

High-throughput identification of immunoreactive peptides and corresponding proteins from Anaplasma platys and Ehrlichia canis using peptide microarray chips

Llanes, Alejandro; Madesh, Swetha; Brangulis, Kalvis; Rajeev, Sreekumari
Front Cell Infect Microbiol.
Jan 2025
10.3389/fcimb.2025.1671309
INTRODUCTION: Anaplasma platys and Ehrlichia canis are rickettsial pathogens infecting dogs, with a worldwide distribution. Both species are obligate intracellular pathogens and colonize bone marrow-derived cells, with coinfections frequently reported in dogs. Although E. canis immunodominant proteins have been thoroughly characterized, very few high-throughput studies have been conducted to identify immunogenic proteins from Anaplasma spp. In this study, we used a methodology based on peptide microarray chips to identify immunoreactive peptides, either shared or species-specific, in the complete theoretical proteomes of both pathogens. METHODS: B-cell epitopes were predicted in the corresponding proteins from both species and ranked for synthesis on the peptide microarrays. These microarrays were screened with serum samples from antibody-positive dogs, as well as negative control sera from unexposed dogs. Additionally, we assessed the feasibility of integrating evidence gathered at the level of individual peptides to identify potentially immunogenic proteins contributing to the patterns of immunoreactivity observed on microarrays. RESULTS: Screening of peptide microarrays resulted in complex antibody reactivity patterns against thousands of peptides. After discarding peptides with cross-reactivity to negative control sera, we identified over 1,200 immunoreactive peptides, including ~80 peptides shared between the two species with almost identical sequences. Despite screening linear peptides, we were able to identify proteins previously reported as immunodominant in E. canis, some of which contain predominantly conformational epitopes. DISCUSSION: Our results suggest that a high-throughput strategy based on peptide microarrays is an effective approach for the rapid identification of immunoreactive peptides and the underlying immunogenic proteins. This study provides a foundation for developing novel diagnostic tools and vaccine candidates against A. platys and E. canis, including potential combined or multivalent formulations targeting both pathogens.

A Quantum Vaccinomics Approach for the Design and Production of MSP4 Chimeric Antigen for the Control of Anaplasma phagocytophilum Infections

de la Fuente, José; Moraga-Fernández, Alberto; Alberdi, Pilar; Díaz-Sánchez, Sandra; García-Álvarez, Olga; Fernández-Melgar, Rubén; Contreras, Marinela
Vaccines.
Nov 2022
10.3390/vaccines10121995
Anaplasma phagocytophilum Major surface protein 4 (MSP4) plays a role during infection and multiplication in host neutrophils and tick vector cells. Recently, vaccination trials with the A. phagocytophilum antigen MSP4 in sheep showed only partial protection against pathogen infection. However, in rabbits immunized with MSP4, this recombinant antigen was protective. Differences between rabbit and sheep antibody responses are probably associated with the recognition of non-protective epitopes by IgG of immunized lambs. To address this question, we applied quantum vaccinomics to identify and characterize MSP4 protective epitopes by a microarray epitope mapping using sera from vaccinated rabbits and sheep. The identified candidate protective epitopes or immunological quantum were used for the design and production of a chimeric protective antigen. Inhibition assays of A. phagocytophilum infection in human HL60 and Ixodes scapularis tick ISE6 cells evidenced protection by IgG from sheep and rabbits immunized with the chimeric antigen. These results supported that the design of new chimeric candidate protective antigens using quantum vaccinomics to improve the protective capacity of antigens in multiple hosts.

In silico and in vitro arboviral MHC class I-restricted-epitope signatures reveal immunodominance and poor overlapping patterns

Lopes-Ribeiro, Ágata; Araujo, Franklin Pereira; Oliveira, Patrícia de Melo; Teixeira, Lorena de Almeida; Ferreira, Geovane Marques; Lourenço, Alice Aparecida; Dias, Laura Cardoso Corrêa; Teixeira, Caio Wilker; Retes, Henrique Morais; Lopes, Élisson Nogueira; Versiani, Alice Freitas; Barbosa-Stancioli, Edel Figueiredo; da Fonseca, Flávio Guimarães; Martins-Filho, Olindo Assis; Tsuji, Moriya; Peruhype-Magalhães, Vanessa; Coelho-dos-Reis, Jordana Grazziela Alves
Front. Immunol..
Nov 2022
10.3389/fimmu.2022.1035515
Introduction The present work sought to identify MHC-I-restricted peptide signatures for arbovirus using in silico and in vitro peptide microarray tools. Methods First, an in-silico analysis of immunogenic epitopes restricted to four of the most prevalent human MHC class-I was performed by identification of MHC affinity score. For that, more than 10,000 peptide sequences from 5 Arbovirus and 8 different viral serotypes, namely Zika (ZIKV), Dengue (DENV serotypes 1-4), Chikungunya (CHIKV), Mayaro (MAYV) and Oropouche (OROV) viruses, in addition to YFV were analyzed. Haplotype HLA-A*02.01 was the dominant human MHC for all arboviruses. Over one thousand HLA-A2 immunogenic peptides were employed to build a comprehensive identity matrix. Intending to assess HLAA*02:01 reactivity of peptides in vitro, a peptide microarray was designed and generated using a dimeric protein containing HLA-A*02:01. Results The comprehensive identity matrix allowed the identification of only three overlapping peptides between two or more flavivirus sequences, suggesting poor overlapping of virus-specific immunogenic peptides amongst arborviruses. Global analysis of the fluorescence intensity for peptide-HLA-A*02:01 binding indicated a dose-dependent effect in the array. Considering all assessed arboviruses, the number of DENV-derived peptides with HLA-A*02:01 reactivity was the highest. Furthermore, a lower number of YFV-17DD overlapping peptides presented reactivity when compared to non-overlapping peptides. In addition, the assessment of HLA-A*02:01-reactive peptides across virus polyproteins highlighted non-structural proteins as “hot-spots”. Data analysis supported these findings showing the presence of major hydrophobic sites in the final segment of non-structural protein 1 throughout 2a (Ns2a) and in nonstructural proteins 2b (Ns2b), 4a (Ns4a) and 4b (Ns4b). Discussion To our knowledge, these results provide the most comprehensive and detailed snapshot of the immunodominant peptide signature for arbovirus with MHC-class I restriction, which may bring insight into the design of future virus-specific vaccines to arboviruses and for vaccination protocols in highly endemic areas.

Cyclic constrained immunoreactive peptides from crucial P. falciparum proteins: potential implications in malaria diagnostics

Vashisht, Kapil; Srivastava, Sukrit; Vandana, Vandana; Das, Ram; Sharma, Supriya; Bhardwaj, Nitin; Anvikar, Anupkumar R; Singh, Susheel Kumar; Kim, Tong-Soo; Na, Byoung-Kuk; Shin, Ho-Joon; Pandey, Kailash C.
Translational Research.
Nov 2022
10.1016/j.trsl.2022.06.008
Malaria is still a global challenge with significant morbidity and mortality, especially in the African, South-East Asian, and Latin American regions. Malaria diagnosis is a crucial pillar in the control and elimination efforts, often accomplished by the administration of mass-scale Rapid diagnostic tests (RDTs). The inherent limitations of RDTs- insensitivity in scenarios of low transmission settings and deletion of one of the target proteins- Histidine rich protein 2/3 (HRP-2/3) are evident from multiple reports, thus necessitating the need to explore novel diagnostic tools/targets. The present study used peptide microarray to screen potential epitopes from 13 antigenic proteins (CSP, EXP1, LSA1, TRAP, AARP, AMA1, GLURP, MSP1, MSP2, MSP3, MSP4, P48/45, HAP2) of P. falciparum. Three cyclic constrained immunoreactive peptides- C6 (EXP1), A8 (MSP2), B7 (GLURP) were identified from 5458 cyclic constrained peptides (in duplicate) against P. falciparum-infected sera. Peptides (C6, A8, B7- cyclic constrained) and (G11, DSQ, NQN- corresponding linear peptides) were fairly immunoreactive towards P. falciparum-infected sera in dot-blot assay. Using direct ELISA, cyclic constrained peptides (C6 and B7) were found to be specific to P. falciparum-infected sera. A substantial number of samples were tested and the peptides successfully differentiated the P. falciparum positive and negative samples with high confidence. In conclusion, the study identified 3 cyclic constrained immunoreactive peptides (C6, B7, and A8) from P. falciparum secretory/surface proteins and further validated for diagnostic potential of 2 peptides (C6 and B7) with field-collected P. falciparum-infected sera samples.

Differential patterns of cross-reactive antibody response against SARS-CoV-2 spike protein detected for chronically ill and healthy COVID-19 naïve individuals

Jaago, Mariliis; Rähni, Annika; Pupina, Nadežda; Pihlak, Arno; Sadam, Helle; Tuvikene, Jürgen; Avarlaid, Annela; Planken, Anu; Planken, Margus; Haring, Liina; Vasar, Eero; Baćević, Miljana; Lambert, France; Kalso, Eija; Pussinen, Pirkko; Tienari, Pentti J.; Vaheri, Antti; Lindholm, Dan; Timmusk, Tõnis; Ghaemmaghami, Amir M.; Palm, Kaia
Sci Rep.
Oct 2022
10.1038/s41598-022-20849-6
Immunity to previously encountered viruses can alter response to unrelated pathogens. We reasoned that similar mechanism may also involve SARS-CoV-2 and thereby affect the specificity and the quality of the immune response against the virus. Here, we employed high-throughput next generation phage display method to explore the link between antibody immune response to previously encountered antigens and spike (S) glycoprotein. By profiling the antibody response in COVID-19 naïve individuals with a diverse clinical history (including cardiovascular, neurological, or oncological diseases), we identified 15 highly antigenic epitopes on spike protein that showed cross-reactivity with antigens of seasonal, persistent, latent or chronic infections from common human viruses. We observed varying degrees of cross-reactivity of different viral antigens with S in an epitope-specific manner. The data show that pre-existing SARS-CoV-2 S1 and S2 cross-reactive serum antibody is readily detectable in pre-pandemic cohort. In the severe COVID-19 cases, we found differential antibody response to the 15 defined antigenic and cross-reactive epitopes on spike. We also noted that despite the high mutation rates of Omicron (B.1.1.529) variants of SARS-CoV-2, some of the epitopes overlapped with the described mutations. Finally, we propose that the resolved epitopes on spike if targeted by re-called antibody response from SARS-CoV-2 infections or vaccinations can function in chronically ill COVID-19 naïve/unvaccinated individuals as immunogenic targets to boost antibodies augmenting the chronic conditions. Understanding the relationships between prior antigen exposure at the antibody epitope level and the immune response to subsequent infections with viruses from a different strain is paramount to guiding strategies to exit the COVID-19 pandemic.

Vivaxin genes encode highly immunogenic, non-variant antigens on the Trypanosoma vivax cell-surface

Romero-Ramirez, Alessandra; Casas-Sánchez, Aitor; Autheman, Delphine; Duffy, Craig W.; Brandt, Cordelia; Clare, Simon; Harcourt, Katherine; André, Marcos Rogério; de Almeida Castilho Neto, Kayo José Garcia; Teixeira, Marta M. G.; Machado, Rosangela Zacharias; Coombes, Janine; Flynn, Robin J.; Wright, Gavin J.; Jackson, Andrew P.
PLoS Negl Trop Dis.
Sep 2022
10.1371/journal.pntd.0010791
Trypanosoma vivax is a unicellular hemoparasite, and a principal cause of animal African trypanosomiasis (AAT), a vector-borne and potentially fatal livestock disease across sub-Saharan Africa. Previously, we identified diverse T. vivax-specific genes that were predicted to encode cell surface proteins. Here, we examine the immune responses of naturally and experimentally infected hosts to these unique parasite antigens, to identify immunogens that could become vaccine candidates. Immunoprofiling of host serum shows that one particular family (Fam34) elicits a consistent IgG antibody response. This gene family, which we now call Vivaxin, encodes at least 124 transmembrane glycoproteins that display quite distinct expression profiles and patterns of genetic variation. We focused on one gene (viv-β8) that encodes one particularly immunogenic vivaxin protein and which is highly expressed during infections but displays minimal polymorphism across the parasite population. Vaccination of mice with VIVβ8 adjuvanted with Quil-A elicits a strong, balanced immune response and delays parasite proliferation in some animals but, ultimately, it does not prevent disease. Although VIVβ8 is localized across the cell body and flagellar membrane, live immunostaining indicates that VIVβ8 is largely inaccessible to antibody in vivo. However, our phylogenetic analysis shows that vivaxin includes other antigens shown recently to induce immunity against T. vivax. Thus, the introduction of vivaxin represents an important advance in our understanding of the T. vivax cell surface. Besides being a source of proven and promising vaccine antigens, the gene family is clearly an important component of the parasite glycocalyx, with potential to influence host-parasite interactions.

Anti-neuronal antibodies against brainstem antigens are associated with COVID-19

Lucchese, Guglielmo; Vogelgesang, Antje; Boesl, Fabian; Raafat, Dina; Holtfreter, Silva; Bröker, Barbara M.; Stufano, Angela; Fleischmann, Robert; Prüss, Harald; Franke, Christiana; Flöel, Agnes
eBioMedicine.
Sep 2022
10.1016/j.ebiom.2022.104211
Background Understanding how SARS-CoV-2 affects respiratory centres in the brainstem may help to preclude assisted ventilation for patients in intensive care setting. Viral invasion appears unlikely, although autoimmunity has been implicated, the responsible antigens remain unknown. We previously predicted the involvement of three epitopes within distinct brainstem proteins: disabled homolog 1 (DAB1), apoptosis-inducing-factor-1 (AIFM1), and surfeit-locus-protein-1 (SURF1). Methods Here, we used microarrays to screen serum from COVID-19 patients admitted to intensive care and compared those with controls who experienced mild course of the disease. Findings The results confirm the occurrence of IgG and IgM antibodies against the hypothesised epitopes in COVID-19 patients. Importantly, while IgM levels were similar in both groups, IgG levels were significantly elevated in severely ill patients compared to controls, suggesting a pathogenic role of IgG. Interpretation The newly discovered anti-neuronal antibodies might be promising markers of severe disease and the targeted peptide epitopes might be used for targeted immunomodulation. Further work is needed to determine whether these antibodies may play a role in long-COVID.

Identification of Equine Arteritis Virus Immunodominant Epitopes Using a Peptide Microarray

Mayers, Jo; Westcott, David; Steinbach, Falko
Viruses.
Aug 2022
10.3390/v14091880
Using the commercially available PEPperCHIP® microarray platform, a peptide microarray was developed to identify immunodominant epitopes for the detection of antibodies against Equine arteritis virus (EAV). For this purpose, the whole EAV Bucyrus sequence was used to design a total of 1250 peptides that were synthesized and spotted onto a microarray slide. A panel of 28 serum samples representing a selection of EAV strains was tested using the microarray. Of the 1250 peptides, 97 peptides (7.76%) showed reactivity with the EAV-positive samples. No single peptide was detected by all the positive serum samples. Seven peptides repeatedly showed reactivity above the cut-off and were considered to have diagnostic potential. Five of these peptides were within the immunodominant GP5 protein and two were within the replicase polyprotein regions NSP2 and NSP10, located in ORF1. The diagnostic sensitivity of the seven peptides selected was low, ranging from 5% to 55%; however, the combined diagnostic sensitivity and specificity of the seven peptides was 90% and 100%, respectively. This data demonstrate that multiple peptide sequences would be required to design a comprehensive serological test to cover the diversity of the EAV strains and the individual immune responses of horses.

Mapping and Validation of Peptides Differentially Recognized by Antibodies from the Serum of Yellow Fever Virus-Infected or 17DD-Vaccinated Patients

Oliveira, Eneida Santos; Tavares, Naiara Clemente; Colombarolli, Stella Garcia; Batista, Izabella Cristina Andrade; Nascimento, Camila Sales; Felgner, Philip Louis; de Assis, Rafael Ramiro; Calzavara-Silva, Carlos Eduardo
Viruses.
Jul 2022
10.3390/v14081645
Yellow Fever disease is caused by the Yellow Fever virus (YFV), an arbovirus from the Flaviviridae family. The re-emergence of Yellow Fever (YF) was facilitated by the increasing urbanization of sylvatic areas, the wide distribution of the mosquito vector, and the low percentage of people immunized in the Americas, which caused severe outbreaks in recent years, with a high mortality rate. Therefore, serological approaches capable of discerning antibodies generated from the wild-type (YFV-WT) strain between the vaccinal strain (YFV-17DD) could facilitate vaccine coverage surveillance, enabling the development of strategies to avoid new outbreaks. In this study, peptides were designed and subjected to microarray procedures with sera collected from individuals infected by WT-YFV and 17DD–YFV of YFV during the Brazilian outbreak of YFV in 2017/2018. From 222 screened peptides, around ten could potentially integrate serological approaches aiming to differentiate vaccinated individuals from naturally infected individuals. Among those peptides, one was synthesized and validated through ELISA.

Quote form