PepMix™
The use of complete protein-spanning mixtures of overlapping peptides is extremely efficient for immunostimulation of T-lymphocytes and diagnostic applications. JPT Peptide Technologies offers cost-effective ready-to-use PepMixes™ on a per-test basis. In addition, customized PepMixes™ will be prepared in a fast and economical manner according to the specific needs of the customer.
In order to establish reliable and validated T-cell assays such as ELISPOT, appropriate positive and negative controls are essential. Selection of such controls helps to confirm proper functionality of the assay as well as viability of the cells used. In addition, high quality controls are most important to establish standardized assay protocols in clinical setups. Compared to commonly used controls like phytohemagglutinin (PHA) and concanavalin A (ConA) that stimulate T-cells as well as other cell types or full length antigens, synthetic peptide pools offer the advantage of a high batch-to-batch reproducibility, applicability of reliable chemical and biochemical QC/QA measures and shelf stability when stored as freeze dried aliquots.
JPT offers several synthetic peptide pools that can serve as positive controls by stimulating antigen specific CD8+ T cell responses (CEF pools) or both CD4+ & CD8+ T-cell responses (HCMV pp65, IE-1, IE-2 pools). Whereas our CEF pools contain 23 (standard) or 32 (extended) different class-I restricted peptides deducted from CMC, EBV and Influenca virus, our CMV peptide pools are constructed from 15meric peptide scans overlapping by 11 amino acids through full length antigens. Thus, the latter pools are able to stimulate both CD4+ and CD8+ responses within a single control experiment.
In addition, JPT offers a variety of peptide pool based negative controls that offer a complement to the commonly used protocols that make use of cells in medium without respective antigen stimulation.
PepMixes™ of the following proteins are available in stock:
Positive control pools for T-Cell Assays
| PepMix | Product Code | Protein | Source | Primary accession number |
Peptides |
| HCMVA (IE-1) | PM-IE1 | 55 kDa immediate-early protein 1 (VIE1_HCMVA) | Human cytomegalovirus | P13202 | 120 |
| HCMVA (IE-2) | PM-IE2 | 45 kDa immediate-early protein 2 (VIE2_HCMVA) | Human cytomegalovirus | P19893 | 143 |
| HCMVA (pp65) | PM-PP65 | 65 kDa lower matrix phosphoprotein (PP65_HCMVA) | Human cytomegalovirus | P06725 | 138 |
| CEF Pool (standard) | PM-CEF-S | defined HLA class I-restricted T-cell epitopes |
Human cytomegalovirus, Epstein-Barr virus, Influenza virus |
n.a. | 23 |
| CEF Pool (extended) | PM-CEF-E | defined HLA class I-restricted T-cell epitopes |
Human cytomegalovirus, Epstein-Barr virus, Influenza virus |
n.a. | 32 |
Negative control pools for T-Cell Assays
| PepMix | Product Code | Protein | Source | Primary accession number |
Peptides |
| ACTS_Human | PM-ACTS | Alpha-actin-1 | Homo Sapiens | P68133 | 92 |
| HLA classI Ig-like C1 domain |
PM-C1 | Human | HLA | n.a. | 19 |
| HIV (Con B gag motif) | PM-HIVgag | Con B gag motif | Human immunodefficiency virus | n.a. | 123 |
| MOG Human (aa 30 -154) |
PM-MOG | Myelin-Oligo dentrocyte glycoprotein |
Homo Sapiens | Q16653 | 29 |
Infectious Diseases
| PepMix | Product Code | Protein | Source | Primary accession number |
Peptides |
| BKV (capsid protein VP1) |
PM-BKV-VP1 | Capsid protein VP1 (VP1_POVBA) | BK polyomavirus | P14996 | 88 |
| BKV (capsid protein VP2) |
PM-BKV-VP1 | Capsid protein VP2 (VP2_POVBK) | BK polyomavirus | P03094 | 85 |
| BKV (VP3) | PM-BKV-VP3 | Capsid protein VP2 (VP3 isoform) | BK polyomavirus | P03094-2 | 56 |
| BKV (large T antigen) |
PM-BKV-LTA | Large T antigen (LT1_POVBA) | BK polyomavirus | P14999 | 170 |
| BKV (small T antigen) |
PM-BKV-StA | Small T antigen (ST1_POVBK) | BK polyomavirus | P03082 | 41 |
| Candida (MP65) | PM-CAND-MP65 | Mannoprotein MP | Candida albicans | Q9HEP1 | 92 |
| EBV (BZLF1) | PM-EBV-BZLF1 | Trans-activator protein BZLF1 (BZLF1_EBV) | Epstein-Barr virus | P03206 | 59 |
| EBV (EBNA1) | PM-EBV-EBNA1 | Epstein-Barr virus Nuclear Antigen | Epstein-Barr virus | YP401677 | 158 |
| EBV (EBNA3a) | PM-EBV-EBNA3a | Epstein-Barr virus Nuclear Antigen | Epstein-Barr virus | YP_401669 | 234 |
| EBV (EBNA3b) | PM-EBV-EBNA3b | Epstein-Barr virus Nuclear Antigen | Epstein-Barr virus | ABB89244 | 279 |
| EBV (EBNA3c) | PM-EBV-EBNA3c | Epstein-Barr virus Nuclear Antigen | Epstein-Barr virus | ABB89245 | 265 |
| EBV (LMP1) | PM-EBV-LMP1 | Latent membrane protein 1 (LMP1_EBV) | Epstein-Barr virus | P03230 | 94 |
| EBV (LMP2) | PM-EBV-LMP2 | Latent membrane protein 2 (LMP2_EBV) | Epstein-Barr virus | P13285 | 122 |
| HAdV-3 (hexon protein) |
PM-HadV3 | Hexon protein (HEX_ADE03) | Human adenovirus 3 | P36849 | 234 |
| HAdV-5 (penton protein) |
PM-HadV5 | Penton protein (PEN3_ADE05) | Human adenovirus 5 | P12538 | 140 |
| HBV (large env.protein) |
PM-HBV-lEV | Large surface protein | Hepatitis B virus | P17101 | 98 |
| HCMVA (IE-1) | PM-IE1 | 55 kDa immediate-early protein 1 (VIE1_HCMVA) | Human cytomegalovirus | P13202 | 120 |
| HCMVA (IE-2) | PM-IE2 | 45 kDa immediate-early protein 2 (VIE2_HCMVA) | Human cytomegalovirus | P19893 | 143 |
| HCMVA (pp65) | PM-pp65 | 65 kDa lower matrix phosphoprotein (PP65_HCMVA) | Human cytomegalovirus | P06725 | 138 |
| HCMVA (UL40) | PM-UL40 | Uncharacterized protein UL40 (UL40_HCMVA) | Human cytomegalovirus | P16780 | 53 |
| HHV6 U54 | PM-HHV-U54 | U54 | Human Herpesvirus 6B | Q9QJ29 | 112 |
| HIV-1 (Con B gag motif) |
PM-HIVgag | Con B gag motif | Human immunodeficiency virus |
n.a. | 123 |
| Influenza A (MP1 protein) |
PM-INFA-MP1 | Membrane protein M1 | Influenza A virus | AAA43277 | 110 |
| Influenza A (NP protein) |
PM-INFA-NP | Nucleocapsid protein | Influenza A virus | ABB79814 | 61 |
| RSV (nucleocapsid protein N) | PM-RSV-NCPN | Nucleocapsid protein | Human respiratory syncytial virus | AAB82431 | 95 |
| RSV (protein F) | PM-RSV-FGF0 | Fusion glycoprotein F0 | Human respiratory syncytial virus | 036634 | 141 |
| Yellow fever NS4B | PM-YF-NS4B | Genome polyprotein | Yellow fever virus | PO3314 | 60 |
Cancer
| PepMix | Product Code | Protein | Source | Primary accession number |
Peptides |
| CEA | PM-CEA | Carcinoembryonic antigen | Homo sapiens | P06731 | 173 |
| Claudin-6 | PM-Cl-6 | Claudin-6 | Homo Sapiens | P56747 | 53 |
| c-Myc (MYC_Human) | PM-cMyc | Myc proto-oncogene | Homo sapiens | P01106 | 107 |
| Cyclin B1 | PM-CYC-B1 | CCNB1 | Homo sapiens | P14635 | 106 |
| ERBB2_ECD | PM-ERB_ECD | ERBB2 (-1 1-683) | Homo sapiens | P04626 | 168 |
| HER 2 ICD | PM-ERB-ICD | ERBB2 (-1 662-1255) | Homo sapiens | P04626 | 146 |
| Histone H1.2 | PM-Histone H1 | HIST1H1C | Homo sapiens | P16403 | 51 |
| Histone H4_Human | PM-H4 | Histone H4 | Homo sapiens | P62805 | 23 |
| Melan-A/MART-1 | PM-MelA | MELAN-A protein (MAR1_HUMAN) | Homo sapiens | Q16655 | 27 |
| MAGEA1 | PM-MAGEA1 | Melanoma-associated antigen 1 | Homo sapiens | P43355 | 75 |
| MAGEA3 | PM-MAGEA3 | Melanoma-associated antigen 3 | Homo sapiens | P43357 | 76 |
| MAGEA4 | PM-MAGEA4 | Melanoma antigen family A, 4 | Homo sapiens | P43358 | 77 |
| Mammaglobin A | PM-MamA | Mammaglobin A (SG2A2_human) | Homo sapiens | Q13296 | 21 |
| MMP11 | PM-MMP11 | Stromelysin 3 | Homo sapiens | P24347 | 121 |
| MVA 018L | PM-MVA_018L | Host range protein 2 | Vaccinia virus | P68598 | 35 |
| MVA 074R | PM-MVA-074R | Putative 49.8k protein | Vaccinia virus | O57196 | 106 |
| MVA 093L | PM-MVA-093L | IMV associated protein p35 | Vaccinia virus | O57206 | 79 |
| MVA 105L | PM-MVA-105L | Cell-surface binding protein | Vaccinia virus | O57211 | 74 |
| MVA 121L (1+2) | PM-MVA-121L | Major Core protein p4a | Vaccinia virus | O57223 | 111+112 |
| MVA 189R | PM-MVA-189R | Putative 21.7k protein | Vaccinia virus | O57265 | 45 |
| NY-ESO-1 | PM-NYE | Cancer/testis antigen 1 (CTG1B_HUMAN) | Homo sapiens | P78358 | 43 |
| P53_human | PM-p53 | Cellular Tumor antigen p53 | Homo sapiens | P04637 | 96 |
| PRAME/OIP4 | PM-OIP4 | Melanoma antigen preferentially expressed in tumors (MAPE_HUMAN) |
Homo sapiens | P78395 | 125 |
| PSA (human) | PM-PSA | Prostate-specific antigen | Homo sapiens | P07288 | 63 |
| SOX-2 | PM-SOX-2 | Transcription factor SOX-2 | Homo sapiens | P48431 | 77 |
| SSX2 | PM-SSX2 | SSX2 | Homo sapiens | Q16385 | 45 |
| Survivin_BIRC5 | PM-Survivin | Baculoviral IAP repeat-containing protein 5 |
Pongo abelii (Sumatran orangutan) |
Q5RAH9 | 33 |
| TARP | PM-TARP | TCR gamma alternate reading frame protein isoform 1 |
Homo sapiens | NP_001003799 | 12 |
| TRP 2 | PM-TRP-2 | L-dopachrome tautomerase | Homo sapiens | P40126 | 127 |
| Tyrosinase_human | PM-Tyr_H | Tyrosinase | Homo sapiens | P14679 | 117 |
| Tyrosinase_mouse | PM-Tyr_M | Tyrosinase | Mus musculus (mouse) | P11344 | 117 |
| WRKY 47 | PM-WRK | Probable WKRY transcription factor 47 | Arabidopsis thaliana | Q9ZS17 | 120 |
| WT1 (WT33) | PM-WT1 | Wilms tumor protein (WT1_HUMAN) | Homo sapiens | P19544 | 110 |
Custom PepMixes™
Custom PepMixes are available via our High Throughput Custom Synthesis program. Protein primary sequences provided will be chemically synthesized as overlapping peptides, purified and analyzed to comply with the requirements of T-cell assays. All production steps are optimized to avoid contaminations and failure sequences which may lead to false positive T-cell responses or inhibition of T-cell responses.
Pooling and aliquotation will be performed using validated protocols ensuring the presence of all peptides in the respective pools. Our fully automated aliquotation service yields freeze dried pool aliquots avoiding deterioration of peptides in DMSO stock solutions.
Applications for PepMix™
- Immunostimulation of T-lymphocytes
- Monitoring of immune status during diseases
- Assessment of vaccine efficacy
Benefits of PepMix™
- Equivalent or better stimulation of CD4 and CD8
- T-cell responses compared to whole protein antigens
- Simultaneous detection of CD4 and CD8 responses in a single sample
- Improved responses in stored blood and PBM cells compared to whole protein antigens
Single Peptides
In addition to the mixes, peptides can also be ordered as single peptide aliquots to validate the results found using the mixes. All peptides and mixes will be quality controlled to ensure batch-to-batch reproducibility!
Testimonials
"I have used JPT's peptide pools (PepMix) for years now, with great satisfaction! First CMV derived peptides and now also EBV. So far we have published data in one publication and plan several more in the near future."
Anna Karin Lidehaell (Uppsala University, Clin. Immunol., Uppsala, Sweden)
"The focus of the Clinical Immunology Group at the German Rheumatism Research Centre is to study the role of cells and molecules in the origin and progression of autoimmune and other diseases. In our hands PepMixes™ were found to be effective in monitoring the immune status of various patient populations but show also promise for the development of novel immuno therapy approaches."
Andreas Thiel, PhD (German Rheumatism Research Centre, Clin. Immunol., Berlin, Germany)
ZellNet Consulting - The Art of the Elispot Technique recommends JPT's PepMixes.
Selected References:
- Clonotype Analysis of Cytomegalovirus-Specific Cytotoxic T Lymphocytes
Babel et al., Journal of the American Society of Nephrology (2008) (abstract) - Prophylactic Transfer of CD8-Depleted Donor Lymphocytes after T-cell Depleted Reduced-Intensity Transplantation
Meyer et al., Blood (2008) (abstract) - Massive Load of Functional Effector CD4+ and CD8+ T Cells against Cytomegalovirus in Very Old Subjects
Myers et al., J. Immunol. (2007) (abstract) - Reconstitution of Adenovirus-Specific Cell-mediated immunity in pediatric patients after Hematopoietic Stem Cell Transplantation
Myers et al., Bone Marrow Transplant. (2007) (abstract) - Monoculture-Derived T Lymphocytes Specific for Multiple Viruses Expand and Produce Clinically Relevant Effects in Immunocompromised Individuals
Leen at al., Nat. Med. (2006) (abstract) - Results of a Cytomegalovirus (CMV)-Specific CD8+/Interferon- Gamma+ Cytokine Flow Cytometry Assay Correlate with Clinical Evidence of Protective Immunity in Patients with AIDS with CMV Retinitis
Jacobson et al., J. Infect. Dis. (2004) (abstract) - Functional Comparison of T Cells Recognizing Cytomegalovirus pp65 and Intermediate-Early Antigen Polypeptides in Hematopoietic Stem-Cell Transplant and Solid Organ Transplant Recipients
Lacey et al., J. Infect. Dis. (2006) (abstract) - Acquisition of Direct Antiviral Effector Functions by CMV-Specific CD4+ T Lymphocytes with Cellular Maturation
Casazza et al., J. Exp. Med. (2006) (abstract) - Protection from Cytomegalovirus After Transplantation is Correlated with Immediate Early 1–specific CD8 T cells
Bunde et al., Eur. J. Exp. Med. (2005) (abstract) - HLA Type-Independent Generation of Antigen-Specific T Cells for Adoptive Immunotherapy
Hammer et al., Eur. J. Immunol. (2005) (abstract) - Generation of Cytomegalovirus (CMV)-Specific T-lymphocytes Using Protein-Spanning Pools of pp65-Derived Overlapping Pentadecapeptides for Adoptive Immunotherapy
Trivedi et al., Blood (2005) (abstract) - Confirmation of Mycobacterium Tubercolosis Infection by Flow Cytometry After Ex Vivo Incubation of Peripheral Blood T Cells with an ESAT-6-Derived Peptide Pool
Tesfa et al., Cytometry Part B (Clinical Cytometry) (2004) (abstract) - CMV Antigen-Specific CD4+ and CD8+ T Cell IFN Expression and Proliferation Responses in Healthy CMV-Seropositive Individuals
Sinclair et al., Viral Immunol. (2004), (abstract) - Recent Advances in the Development of HIV-1 Vaccines Using Replication-Incompetent Adenovirus Vectors
Shiver and Emini, Annual Review of Medicine (2004), (abstract) - Mapping T Cell Epitopes by Flow Cytometry
Hoffmeister et al., Methods (2003), (abstract) - Cytomegalovirus (CMV) Phosphoprotein 65 Makes a Large Contribution to Shaping the T Cell Repertoire in CMV-Exposed Individuals
Kern et al., J. Infect. Dis.(2002), (abstract) - Use of Overlapping Peptide Mixtures as Antigens for Cytokine Flow Cytometry
Maecker et al., J. Immunol. Methods (2001), (abstract) - Analysis of CD8 T Cell Reactivity to Cytomegalovirus Using Protein-Spanning Pools of Overlapping Pentadecapeptides
Kern et al., Eur. J. Immunol. (2000), (abstract) - Putative Immunodominant Human Immunodeficiency Virus-specific CD8+ T-Cell Responses Cannot Be Predicted by Major Histocompatibility Complex Class I Haplotype
Betts et al., J. Virol. (2000), (abstract) - Identification of T-cell Epitopes Using ELISpot and Peptide Pool Arrays, Tobery and Caulfield
Methods in Molecular Medicine : Molecular Diagnosis of Infectious Deseases, 2/e; Edited by J.Decker and U. Reischl; Humana Press Inc, Totowa, NJ, (abstract)
More references under JPT Publications/Literature
