PepTivator SARS-CoV-2 MHC-I 

Two MHC-I specific SARS-CoV-2 PepTivators, consisting of MHC class I-restricted peptides of 8-12 aa length, are available in research grade quality. PepTivator SARS-CoV-2 MHC-I Select covers selected epitopes of the whole proteome of SARS-CoV-2 (SARS-CoV-2 envelope (E) protein, nucleoprotein (N), nucleoprotein (N), ORF1ab, ORF3a, ORF7a, ORF8 and ORF10). In contrast, PepTivator SARS-CoV-2 MHC-I Select Prot_S covers selected epitopes of the SARS-CoV-2 surface or spike glycoprotein (S).

The MHC-I specific SARS-CoV-2 PepTivators can be:

• applied individually to analyze a) CD8+ T cells reactive to the SARS-CoV-2 spike protein (PepTivator SARS-CoV-2 MHC-I Select Prot_S) or b) CD8+ T cells reactive to the whole SARS-CoV-2 proteome without the spike protein (PepTivator SARS-CoV-2 MHC-I Select),
• used together (PepTivator SARS-CoV-2 MHC-I Select plus PepTivator SARS-CoV-2 MHC-I Select Prot_S) to analyze CD8+ T cells reactive to the whole SARS-CoV-2 proteome, or
• used to supplement other PepTivators to boost the CD8+ T cell response and analyze CD4+ as well as CD8+ T cells.

Comparison of T cell reactivity towards the spike protein and other SARS-CoV-2 proteins allows to distinguish vaccine-induced from natural T cell immunity. 
 

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. 

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.
 

PepTivator Peptide Pools consist of mainly of 15-mer peptides with 11-amino-acid overlaps, covering the amino acid sequence of a particular 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.

Our SARS-CoV-2 PepTivator Peptide pools are available in both sizes (6 nmol and 60 nmol per peptide per vial) in research-grade quality. With a broad product selection covering main structural SARS-CoV-2 proteins and an average purity of 70%, they are the perfect choice for your basic research, e.g., to monitor the immune response after natural infections or upon vaccination. 

Highlight: PepTivator SARS-CoV-2 Prot_S Complete is now also available in premium-grade quality. In this specific case individual peptides have a purity of >90%.
 

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

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.   

Since the beginning of the pandemic, the S protein and especially the S1 domain have been considered a potential key target for vaccines, therapeutics, and diagnostic approaches. In fact, thanks to the relentless work of the scientific community, several vaccines based on the spike protein are available today and more are under development.
 

We offer different PepTivator Peptide Pools for the spike protein:

• 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).
• PepTivator SARS-CoV-2 SARS-CoV-2 Prot_S1 spans the complete N-terminal S1 domain of the spike protein (aa 1–692).
• PepTivator SARS-CoV-2 SARS-CoV-2 Prot_S+ spans a part of the C-terminal S2 domain (aa 689–895) and can be combined with SARS-CoV-2 Prot_S to cover the complete S2 domain (and parts of the S1 domain: aa 304–338, 421–475, and 492–519).
• PepTivator SARS-CoV-2 Prot_S Complete covers the complete sequence of the mature spike protein (omitting the first four amino acids of the N-terminal signal peptide). For your convenience, this product is optimized for water solubility, is delivered in one vial, and is available in two different quality grades. The research-grade product has an average purity of 70%, whereas peptides of the premium-grade product are individually purified via HPLC to a purity of >90%.  

To achieve a complete sequence coverage, you can either combine the three PepTivator Peptide Pools (SARS-CoV-2 Prot_S, Prot_S1 and Prot_S+) or choose PepTivator SARS-CoV-2 Prot_S Complete as a convenient all-in-one solution.

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). 
 

Detection of SARS-CoV-2–specific CD3⁺ T cells in individuals recovered from COVID-19

A more detailed analysis of the stimulation of CD4⁺ and CD8⁺ T cells is shown in "exemplary application data" and more examples for the application of SARS-CoV-2 PepTivator Peptide Pools can be found under "references". 

Following ordinary viral evolution, SARS-CoV-2 accumulates mutations over time, leading to new virus variants. With the B.1.1.7 lineage (Alpha variant) and the B.1.351 lineage (Beta variant), it was first reported that mutations have affected immune relevant parts of structural virus proteins. New SARS-CoV-2 lineages are in the spotlight of COVID-19 research as they are able to evade pre-existing antibody responses and appear to be more infectious than the Wuhan wild-type variant.

We offer PepTivator Peptide Pools covering the mutated sequences of the structural proteins of the new SARS-CoV-2 variants (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_S Complete, Prot_N, or Select) to ensure that the immune response towards different virus lineages is detected upon stimulation. Besides the mutation pools, we also offer the respective WT reference pools, covering the homologous sequence domains 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.

We continuously extend our product portfolio as new virus strains arise. The table below gives an overview of available PepTivator Mutation and WT Reference Pools. The products are available in research grade with a size of 6 nmol/peptide.  

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. Therefore it is a key target for vaccines, therapeutics, and diagnostic approaches. With new virus variants emerging, new adapted vaccines were developed based on the S protein of mutated variants e.g. the Omicron variants BA.1 and BA.5. We offer PepTivator Peptide Pools covering the complete coding sequence of the S protein of these variants to facilitate research on immune responses towards these virus variants e.g. in recovered individuals or after vaccination. The products are available in research grade quality with a size of 6 nmol/peptide.

We offer different PepTivator Peptide Pools for the spike protein of mutated variants:

• PepTivator SARS-CoV-2 Prot_S B.1.351/BA.1:

Covers the complete sequence of the mature spike protein (omitting the first four amino acids of the N-terminal signal peptide) of the B.1.1.529/BA.1 lineage (Omicron variant).

• PepTivator SARS-CoV-2 Prot_S B.1.351/BA.5:

Covers the complete sequence of the mature spike protein (omitting the first four amino acids of the N-terminal signal peptide) of the B.1.1.529/BA.5 lineage (Omicron variant).

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

• Aiello, A. et al. (2021) Spike is the most recognized antigen in the whole-blood platform in both acute and convalescent COVID-19 patients. Int. J. Infect. Dis. 106: 338–347.
• Anft, M. et al. (2020) COVID-19 progression is potentially driven by T cell immunopathogenesis. medRxiv 2020.04.28.20083089. Unrefereed preprint.
• Bonifacius, A. et al. (2021) COVID-19 immune signatures reveal stable antiviral T cell function despite declining humoral responses. Immunity 54(2): 340–354.e6.
• 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.
• Demaret, J. et al. (2020) Severe SARS-CoV-2 patients develop a higher specific T cell response. Clin. Transl. Immunology 9(12): e1217.
• Fazolo, T. et al. (2021) Strong anti-viral responses in pediatric COVID-19 patients in South Brazil. medRxiv 2021.04.13.21255139. 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.
• Gil-Manso, S. et al. (2021) ABO blood group is involved in the quality of the specific immune response. bioRxiv 2021.05.23.445114. Unrefereed preprint.
• Grau-Expósito, J. et al. (2020) Peripheral and lung resident T cell responses against SARS-CoV-2. medRxiv 2020.12.02.20238907. Unrefereed preprint.
• Jung, J. H. et al. (2021) SARS-CoV-2–specific T cell memory is sustained in COVID-19 convalescents for 8 months with successful development of stem cell–like memory T cells. medRxiv 2021.03.04.21252658. 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.
• Roukens, A.H.E. et al. (2021) Prolonged activation of nasal immune cell populations and development of tissue-resident SARS-CoV-2–specific CD8 T cell responses following COVID-19. medRxiv 2021.04.19.21255727. 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.
• Shomuradova, A.S. et al. (2020) SARS-CoV-2 epitopes are recognized by a public and diverse repertoire of human T cell receptors. Immunity 53(6): 1245–1257.e5.
• Stephenson, E. et al. (2021) Single-cell multi-omics analysis of the immune response in COVID-19. Nat Med 27: 904–916.
• 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.
• Cotugno, N. et al. (2021) Virological and immunological features of SARS-CoV-2–infected children who develop neutralizing antibodies. Cell Rep. 34(11):108852.
• Meckiff, B. J. et al. (2020) Imbalance of regulatory and cytotoxic SARS-CoV-2–reactive CD4⁺ T cells in COVID-19. 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.
• Riou, C. et al. (2021) Profile of SARS-CoV-2–specific CD4 T cell response: Relationship with disease severity and impact of HIV-1 and active Mycobacterium tuberculosis co-infection. medRxiv 2021.02.16.21251838v1. 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.
• Motozono, C. et al. (2021) An emerging SARS-CoV-2 mutant evading cellular immunity and increasing viral infectivity. bioRxiv 2021.04.02.438288. 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, immunization and vaccination research  

• Au, L. et al. (2021) Cytokine release syndrome in a patient with colorectal cancer after vaccination with BNT162b2. Nat. Med.: Epub ahead of print, May 26.
• Harrington, P. et al. (2021) Single dose of BNT162b2 mRNA vaccine against SARS-CoV-2 induces neutralizing antibody and polyfunctional T cell responses in patients with CML. medRxiv 2021.04.15.21255482. Unrefereed preprint.
• 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.
• Mazzoni, A. et al. (2021) First dose mRNA vaccination is sufficient to reactivate immunological memory to SARS-CoV-2 in ex COVID-19 subjects. medRxiv 2021.03.05.21252590. Unrefereed preprint.
• Nanishi, E. et al. (2021) Alum:CpG adjuvant enables SARS-CoV-2 RBD-induced protection in aged mice and synergistic activation of human elder type 1 immunity. bioRxiv 2021.05.20.444848. Unrefereed preprint.
• Shroff, R.T. et al. (2021) Immune responses to COVID-19 mRNA vaccines in patients with solid tumors on active, immunosuppressive cancer therapy. medRxiv 2021.05.13.21257129. Unrefereed preprint.
• Silva-Cayetano, A. et al. (2021) A booster dose enhances immunogenicity of the COVID-19 vaccine candidate ChAdOx1 nCoV-19 in aged mice. Med (N Y). 2(3): 243–262.e8.
• 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.
• Sir Karakus, G. et al. (2021) Preclinical efficacy and safety analysis of gamma-irradiated inactivated SARS-CoV-2 vaccine candidates. Sci. Rep. 11(1): 5804.
• Ssemaganda A. et al. (2021) Expansion of tissue-resident CD8⁺ T cells and CD4⁺ Th17 cells in the nasal mucosa following mRNA COVID-19 vaccination. bioRxiv 2021.05.07.442971. 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.

Mutated SARS-CoV-2 variants   

• De Marco, L. et al. (2021) Preserved T cell reactivity to the SARS-CoV-2 Omicron variant indicates continued protection in vaccinated individuals. bioRxiv 2021.12.30.474453. 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.
• Geers D. et al. (2021) SARS-CoV-2 variants of concern partially escape humoral but not T cell responses in COVID-19 convalescent donors and vaccines. Sci. Immunol. 6(59): eabj1750.
• Thiruvengadam, R. et al. (2021) Effectiveness of ChAdOx1 nCoV-19 vaccine against SARS-CoV-2 infection during the delta (B.1.617.2) variant surge in India: a test-negative, case-control study and a mechanistic study of post-vaccination immune responses. Lancet Infect. Dis. Epub ahead of print, Nov. 25.

• Untouched isolation of PBMCs from human whole blood: The MACSprep™ PBMC Isolation Kit, human enables a safe and fast untouched isolation of human PBMCs from small whole blood samples without density gradient centrifugation.
• Antigen-specific in vitro stimulation: Our PepTivator Peptide Pools are available for various viruses and enable the stimulation of CD4⁺ and CD8⁺ T cells.
• Enrichment and sorting of antigen- and virus-specific T cells: Antigen- and virus-specific T cells are extremely rare, which makes analysis difficult. Our optimized enrichment and sorting solutions enable you to even analyze the rarest cells. 
  CD137 MicroBead Kit, human and CD154 MicroBead Kit, human for the pre-enrichment of rare virus-reactive T cells based on activation marker expression. 
  IFN-γ Secretion Assay – Cell Enrichment and Detection Kit (PE), human to detect and enrich viable IFN-γ–secreting T cells for any downstream applications. Other specificities are available.
• Multiparameter flow cytometry analysis of antigen- and virus-specific T cells:
  SARS-CoV-2 T Cell Analysis Kits – All-in-one kits for the efficient stimulation and analysis of human SARS-CoV-2–reactive T cells. Kits include all needed buffers and controls, a SARS-CoV-2 PepTivator Peptide Pools of choice and an optimized flow panel.
  REAfinity™ Recombinant Antibodies – Recombinantly engineered for higher purity, superior lot-to-lot consistency, and lowest background. 
  MACSplex Cytokine Kits for a multiplexed analysis of secreted cytokines in cell culture supernatant. 
  Rapid Cytokine Inspector Kits – Unique for the fastest combination of surface marker and intracellular cytokine staining. 
  8-Color Immunophenotyping Kit, anti-human, REAfinity for reliable identification of leukocyte subsets in one simple kit, e.g., monocytes, neutrophils, eosinophils, and T, B, and NK cell populations

Find detailed information on our complete workflow for stimulation, enrichment and analysis of virus-reactive T cells here.