Peptide pools covering the spike glycoprotein: PepTivator SARS‑CoV‑2 Prot_S, Prot_S1, and Prot_S+

The surface or spike glycoprotein (S) plays a critical role in coronavirus infection as it is crucial for recognition and fusion of the virus with the host cell membrane. Two different functional domains of the S protein are responsible for these individual processes: The S1 domain contains the surface binding site to the host cell receptor, the ACE2 receptor, whereas membrane fusion is mediated via the S2 domain. Furthermore, cleavage at the boundary of the S1 and S2 domains (aa residues 685 and 686) via cellular proteases primes the protein for entry into the host cell. Upon fusion of the viral envelope and the host cell membrane, the viral genome enters the host cell and viral replication starts.   

The S protein, and especially the S1 domain, have been considered a potential key target for vaccines, therapeutics, and diagnostic approaches  (PMID: 32106567, 32142651).  

The PepTivator SARS-CoV-2 Prot_S covers selected immunodominant sequence domains of the spike protein (aa 304–338, 421–475, 492–519, 683–707, 741–770, 785–802, and 885–1273; GenBank MN908947.3, Protein QHD43416.1). SARS-CoV-2 Prot_S1 spans the complete N-terminal S1 domain of the spike protein (aa 1–692) and SARS-CoV-2 Prot_S+ a part of the C-terminal S2 domain (aa 689–895). 

All three PepTivator Products can be combined to achieve a complete sequence coverage of the spike protein. Furthermore, the complete S2 domain (and parts of the S1 domain: aa 304–338, 421–475, and 492–519) is covered, when SARS-CoV-2 Prot_S and SARS-CoV-2 Prot_S+ are combined. 

PepTivator SARS‑CoV‑2 Prot_N

Prot_N stands for nucleoprotein (N), also known as nucleocapsid protein. The N protein has multiple functions in the coronavirus replication cycle. Its primary function is viral genome packaging. Furthermore, it is involved in virus transcription and interacts with the viral membrane during virion assembly.  

The N protein has been suggested as a possible target for vaccine development (PMID: 32106567).  

The PepTivator SARS‑CoV‑2 Prot_N covers the complete sequence of the SARS‑CoV‑2 N protein (GenBank MN908947.3, Protein QHD43423.2).    
 

PepTivator SARS‑CoV‑2 Prot_M

Prot_M stands for membrane glycoprotein (M). The M protein is the most abundant structural protein in the coronavirus envelope and thus defines viral morphogenesis. Additionally, the M protein is crucial for efficient virion assembly and involved in viral budding.  

The M protein has been suggested as a possible target for vaccine development (PMID: 16423399).  

The PepTivator SARS‑CoV‑2 Prot_M covers the complete sequence of the SARS‑CoV‑2 M protein (GenBank MN908947.3, Protein QHD43419.1). 
 

All of the above-mentioned PepTivator SARS‑CoV‑2 products are available in two sizes (6 nmol and 60 nmol per peptide per vial) in research-grade quality.
 

NEW: PepTivator SARS-CoV-2 Select

Our PepTivator SARS-CoV-2 Select covers selected immunodominant epitopes of the whole SARS-CoV-2 proteome. The peptides originate from structural proteins (spike protein, membrane protein, nucleoprotein, and envelope protein) as well as non-structural proteins. The included MHC class I– and MHC class II–restricted peptides have a length between 9–22 aa and enable the stimulation of CD4⁺ and CD8⁺ T cells. 

As illustrated by the above-presented data (see “Exemplary application data of SARS‑CoV‑2 PepTivator Peptide Pools”), a strong T cell immune response can be observed upon stimulation with the PepTivator SARS-CoV-2 Select. 

The PepTivator SARS-CoV-2 Select is available in two quality grades:

Premium grade, being perfectly suited for your translational and basic research with a size of 6 nmol/peptide and an overall peptide purity of >80%. The product is manufactured and tested under a quality management system (ISO 9001). 

MACS GMP, supporting your clinical research with a size of 60 nmol/peptide and an overall peptide purity of >90%. The sterile-bottled product (tested according to Ph. Eur.) is manufactured and tested under a quality management system (ISO 13485) and in compliance with relevant GMP guidelines. Designed following the recommendations of USP <1043> on ancillary materials.  
Find out more about our MACS GMP PepTivator Peptide Pools and their application here.
 

Coming soon: PepTivator Peptide Pools covering mutations of new SARS-CoV-2 variants

Following ordinary viral evolution, SARS-CoV-2 accumulates mutations over time, leading to new virus variants. With the recently emerged B.1.1.7 lineage (UK variant), it was first reported that mutations have affected immune relevant parts of structural virus proteins. The B.1.1.7 lineage has been found to be more infectious than former variants. It has become the dominant strain in the UK and first infections with this new variant have been reported in different countries all over the world.

Compared to the Wuhan variant (wild type (WT)) of SARS-CoV-2, the B.1.1.7 lineage, also known as VUI202012/01 or VOC202012/01, displays mutations in the nucleoprotein and spike protein. Soon, we will offer PepTivator Peptide Pools covering these mutated sequences (mutation pools). They can be used to supplement PepTivator Peptide Pools based on the Wuhan variant (PepTivator SARS-CoV-2 Prot_S, Prot_S1, Prot_S+, Prot_N, or Select) to ensure that the immune response towards both virus lineages is detected upon stimulation. Besides the mutation pools, we also offer the respective WT reference pools, including the homologous peptides of the Wuhan variant (WT) without mutation. Thus, comparison of the T cell immune response upon stimulation with the mutation versus reference pools can be used to detect mutation-specific T cell responses. 

B.1.1.7 spike protein 

In total, 10 mutations occurred in the spike protein of the B.1.1.7 lineage (deletion 69, deletion 70, deletion 144, N501Y, A570D, D614G, P681H, T716I, S982A, D1118H) compared to the variant first discovered in Wuhan (WT) (reference genome GenBank MN908947.3). Of particular interest are the N501Y mutation in the receptor binding domain (RBD), as it influences the binding of the virus to the host cell, and the D614G mutation, as it increases the transmissibility of the virus. Our PepTivator SARS-CoV-2 Prot_S B.1.1.7 Mutation Pool contains 34 peptides covering all of the above-mentioned mutations of the spike protein of the B.1.1.7 lineage. PepTivator SARS-CoV-2 Prot_S B.1.1.7 WT Reference Pool consists of the homologous peptides of the Wuhan WT without mutation. 

B.1.1.7 nucleoprotein 

In total, 4 mutations occurred in the nucleoprotein of the B.1.1.7 lineage (D3L, R203K, G204R, S235F) compared to the Wuhan variant (WT) (reference genome GenBank MN908947.3). Our PepTivator SARS-CoV-2 Prot_N B.1.1.7 Mutation Pool contains 9 peptides covering all of the above-mentioned mutations of the nucleoprotein of the B.1.1.7 lineage. PepTivator SARS-CoV-2 Prot_N B.1.1.7 WT Reference Pool consists of the homologous peptides of the Wuhan WT without mutation. 

The products for the mutation and reference pools are available in research grade with a size of 6 nmol/peptide.  
 

Human T cell immunity – CD4⁺ and CD8⁺ T cell response

• Anft, M. et al. (2020) COVID-19 progression is potentially driven by T cell immunopathogenesis. medRxiv 2020.04.28.20083089. Unrefereed preprint.
• Cheung, C. C. L. et al. (2020) Residual SARS-CoV-2 viral antigens detected in gastrointestinal and hepatic tissues from two recovered COVID-19 patients. medRxiv 2020.10.28.20219014. Unrefereed preprint.
• Fendler, A. et al. (2020) Adaptive immunity to SARS-CoV-2 in cancer patients: The CAPTURE study. medRxiv 2020.12.21.20248608. Unrefereed preprint.
• Fischer, D. S. et al. (2020) Single-cell RNA sequencing reveals in vivo signatures of SARS-CoV-2-reactive T cells through ‘reverse phenotyping’. medRxiv 2020.12.07.20245274. Unrefereed preprint.
• Grau-Expósito, J. et al. (2020) Functional profile, homing and residency of protective T cell immune responses against SARS-CoV-2. medRxiv 2020.12.02.20238907. Unrefereed preprint.
• Li, Z. et al. (2020) SARS-CoV-2-specific T cell memory is long-lasting in the majority of convalsecent COVID-19 individuals. bioRxiv 2020.11.15.383463. Unrefereed preprint.
• Schwarzkopf, S. et al. (2021) Cellular immunity in COVID-19 convalescents with PCR-confirmed infection but with undetectable SARS-CoV-2–specific IgG. Emerg. Infect. Dis. 27(1): 122–129.
• Sekine, T. et al. (2020) Robust T cell immunity in convalescent individuals with asymptomatic or mild COVID-19. Cell 183(1): 158–168.e14: Epub ahead of print, Aug. 14.
• Thieme, C. J. et al. (2020) The SARS-CoV-2 T-cell immunity is directed against the spike, membrane, and nucleocapsid protein and associated with COVID-19 severity. medRxiv 2020.05.13.20100636. Unrefereed preprint.
• Vallejo, A. et al. (2020) SARS-CoV-2 cellular immune response in uninfected health care workers with prolonged and close exposure to COVID-19 patients. Research Square rs-55720/v1. Unrefereed preprint.
• Willingham, S. B. et al. (2020) Characterization and phase 1 trial of a B cell activating anti-CD73 antibody for the immunotherapy of COVID-19. medRxiv 2020.09.10.20191486. Unrefereed preprint.

Human CD4⁺ T cell response 

• Bacher, P. et al. (2020) Pre-existing T cell memory as a risk factor for severe 1 COVID-19 in the elderly. medRxiv 2020.09.15.20188896. Unrefereed preprint.
• Meckiff, B. J. et al. (2020) Single-cell transcriptomic analysis of SARS-CoV-2 reactive CD4⁺ T cells. Cell 183(5):1340–1353.e16: Epub ahead of print, Oct. 05.
• Riou, C. et al. (2020) Rapid, simplified whole blood-based multiparameter assay to quantify and phenotype SARS-CoV-2–specific T cells. medRxiv 2020.10.30.20223099. Unrefereed preprint.
• Sattler, A. et al. (2020) SARS–CoV-2–specific T cell responses and correlations with COVID-19 patient predisposition. J. Clin. Invest. 130(12): 6477–6489.

Human CD8⁺ T cell response 

• Habel, J. R. et al. (2020) Suboptimal SARS-CoV-2−specific CD8⁺ T cell response associated with the prominent HLA-A*02:01 phenotype. Proc. Natl. Acad. Sci. USA 117(39): 24384–24391.
• Kusnadi, A. et al. (2020) Severely ill COVID-19 patients display augmented functional properties in SARS-CoV-2-reactive CD8⁺ T cells. bioRxiv 2020.07.09.194027. Unrefereed preprint.

Cell therapy  

• Cooper, R. S. et al. (2021) Rapid GMP-compliant expansion of SARS-CoV-2-specific T cells from convalescent donors for use as an allogeneic cell therapy for COVID-19. Front. Immunol.: Epub ahead of print, Jan. 08.
• Ferreras, C. et al. (2020) SARS-CoV-2 specific memory T lymphocytes from COVID-19 convalescent donors: identification, biobanking and large-scale production for adoptive cell therapy. bioRxiv 2020.10.23.352294. Unrefereed preprint.
• Leung, W. et al. (2020) Successful manufacturing of clinical-grade SARS-CoV-2–specific T cells for adoptive cell therapy. medRxiv 2020.04.24.20077487. Unrefereed preprint.

Animal models and immunization:  

• Haun, B. K. et al. (2020) CoVaccine HT™ adjuvant potentiates robust immune responses to recombinant SARS-CoV-2 Spike S1 immunization. Front. Immunol.: Epub ahead of print, Oct. 30.
• Ishigaki, H. et al. (2021) Neutralizing antibody-dependent and -independent immune responses against SARS-CoV-2 in cynomolgus macaques. Virology 554: 97–105.
• Karakus, G. S. et al. (2020) Preliminary report of preclinical efficacy and safety analysis of gamma-irradiated inactivated SARS-CoV-2 vaccine candidates, SK-01 version 1 and OZG-3861 version 1. bioRxiv 2020.09.04.277426. Unrefereed preprint.
• Singh, D. K. et al. (2020) SARS-CoV-2 infection leads to acute infection with dynamic cellular and inflammatory flux in the lung that varies across nonhuman primate species. bioRxiv 2020.06.05.136481. Unrefereed preprint.
• Watterson, D. et al. (2020) Molecular clamp stabilized Spike protein for protection against SARS-CoV-2. Research Square rs-68892/v1. Unrefereed preprint.

PepTivator® Peptide Pools  consist of short 15-mer peptides with 11-amino-acid overlaps, covering the amino acid sequence of a particular virus-, tumor-, or auto-antigen. 

They bind to MHC class I as well as MHC class II complexes and thus are suitable for the antigen-specific stimulation of CD4⁺ and CD8⁺ T cells. Products containing 6 nmol per peptide are sufficient to stimulate up to 10⁸ cells and products containing 60 nmol per peptide are sufficient to stimulate up to 10⁹ cells.
 

We offer PepTivator Peptide Pools in three quality grades: 

Research grade: Select from a broad range of antigens with an average purity of 70% – the perfect choice for your basic research. 
Premium grade: A selection of individually purified (purity >80%) and endotoxin-tested peptides – perfectly suited for your translational and basic research. 
MACS GMP Grade: Sterile-bottled peptides (tested according to Ph. Eur) with an overall peptide purity of >90%. Manufactured and tested under a quality management system (ISO 13485) and in compliance with relevant GMP guidelines – an excellent fit for the development of clinical applications.     
 

Virus-specific PepTivator Peptide Pools can be used:

• In vaccine research, for example, to analyze T cell immunity after vaccination (PMID: 31137606, 21098232, 30846697).
• For research on dendritic cell vaccination, for example, to pulse antigen-presenting cells (PMID: 30952614).
• To monitor the immune status of patients (PMID: 32169158, 20427631, 26734066, 28960213).

If you are interested in virus-specific PepTivator Products besides SARS-CoV-2, please have a look here.

• Isolation of viable, antigen-specific CD4⁺ T cells with the CD154 MicroBead Kit (see protocol in related documents).
• Isolation of viable, antigen-specific CD4⁺ and CD8⁺ T cells using the MACS® Cytokine Secretion Assay.
• Detection and enrichment of human IFN-γ–secreting cells with Cell Enrichment and Detection Kits.
• Selection of human CD137⁺ cells from PBMCs with the CD137 MicroBead Kit.
• Detection of antigen-specific CD4⁺ and CD8⁺ effector/memory T cells using  MACS® Cytokine Secretion Assays or intracellular cytokine staining.

Find a complete workflow to activate, isolate, and detect virus-specific T cells here.

For the detection of SARS-CoV-2–reactive T cells, we offer an all-in-one kit including a PepTivator Peptide Pool of your choice as well as optimized staining panels and protocol. Get more product information here.