TetTox-B (830-843) MHC II DRB1*07:01
TetTox-B (830-843) DRB1*07:01 is a short part of the tetanus toxin. Tetanus toxin is produced by Clostridium tetani and is a potent neurotoxin. Indeed, tetanus toxin arrives on bloodstream and moves to a receptor complex at the neuromuscular junction.
TetTox-B (830-843) MHC II DRB1*07:01
TetTox-B (830-843) DRB1*07:01 is used to stimulate the immune response and to generate DRB1*07:01-restricted T cells specific for the tetanus toxin peptide. TetTox-B (830-843) is also a classical peptide reference for the binding affinity with DRB1*07:01 molecules.
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We have characterized five human T cell clones specific for tetanus toxin. The combination of different techniques allowed us to precisely map two T cell epitopes within fragments 830-843 and 1273-1284 of tetanus toxin, as formally demonstrated by the use of corresponding synthetic peptides. The three other T cell clones were specific for regions 2-602, 604-742, and 865-1315 of tetanus toxin, respectively. The five T cell clones were shown to be restricted to HLA-DR Ag. Furthermore, the allele of HLA-DR utilized by the various epitopes has been determined. The use of HLA-DR-transfected L cells as APC directly demonstrated that two epitopes, one of which represented by fragment 1273-1284, were recognized in association with HLA-DRw52a. For the other three T cell epitopes, the data strongly suggested they were recognized in association with HLA-DR5. Finally, a sixth T cell clone was shown to be specific for tetanus toxoid, the vaccinal preparation of tetanus toxin, and not for other tetanus toxin fragments. This indicated that immunization with tetanus toxoid probably elicits a T cell response directed only in part against native tetanus toxin.[/et_pb_accordion_item]
2- Demotz S, Matricardi P M, Irle C, Panina P, Lanzavecchia A and Corradin G. J Immunol. 143(12):3881-6 (1989)
Processing of tetanus toxin by human antigen-presenting cells. Evidence for donor and epitope-specific processing pathways
Human T cell clones specific for epitopes 830-843 and 947-967 of tetanus toxin can be differentially activated in vitro when APC (PBL or LCL) from different donors are pulsed with tetanus toxin. Although PBL tested do not seem to exhibit substantial differences in the number of precursor T cells specific for these epitopes, APC from the same donors activate clone KT-2 specific for peptide 830-843 but not clone KT-30 specific for peptide 947-967. These APC express the proper restriction element because they can present the corresponding synthetic peptides. The failure to present a particular epitope might, however, be explained by the absence or presence of a protease(s) required for Ag presentation that may vary for different epitopes. Indeed, the protease inhibitor leupeptin was found to inhibit activation of KT-2 but not KT-30 T cell clone by the KK.35 B cell line normally capable of presenting either epitope. In summary, these data suggest that tetanus toxin processing and epitope formation by APC is distinct in different donors and for different epitopes.
3- Valitutti S, Muller S, Salio M and Lanzavecchia A. J Exp Med. 185(10):1859-64 (1997)
T cell activation by specific antigen results in a rapid and long-lasting downregulation of triggered T cell receptors (TCRs). In this work, we investigated the fate of downregulated TCR- CD3-zeta complexes. T cells stimulated by peptide-pulsed antigen-presenting cells (APCs) undergo an antigen dose-dependent decrease of the total cellular content of TCR-beta, CD3-epsilon, and zeta chains, as detected by FACS(R) analysis on fixed and permeabilized T-APC conjugates and by Western blot analysis on cell lysates. The time course of CD3-zeta chain consumption overlaps with that of TCR downregulation, indicating that internalized TCR-CD3 complexes are promptly degraded. Inhibitors of lysosomal function (bafilomycin A1, folimycin) markedly reduced zeta chain degradation, leading to the accumulation of zeta chain in large Lamp1(+) vesicles. These results indicate that in T cell-APC conjugates, triggered TCRs are rapidly removed from the cell surface and are degraded in the lysosomal compartment.
4- Cancedda C, Suciu-Foca N, Cortesini R and Harris P E. Hum Immunol. 61(6):575-84 (2000)
The aim of our experiments was to determine whether deletion of antigen specific T helper cells could be accomplished by delivering the antigenic peptide to antigen presenting cells. Tetanus toxin peptide residues 830-843 was chosen for these experiments. Two mammalian expression vectors carrying the genes for human Fas ligand and a chimeric invariant chain-tetanus toxin peptide construct were designed. The T cell proliferative response to tetanus toxoid was inhibited when the antigen was presented by autologus monocytes transfected with Fas ligand. T cell mixture experiments using two syngeneic T cell lines specific either for tetanus toxoid or for pertussis toxin demonstrated that the killing effect elicited by the antigen pulsed/Fas ligand-transfected antigen presenting cells was antigen specific. Finally, we demonstrated that transient expression of antigen delivered by plasmid DNA can substitute for soluble antigen in the induction of antigen-specific T cell responses. Antigen presenting cells transfected with the vector carrying Fas ligand and the vector carrying the chimeric invariant chain-peptide antigen gene were shown to inhibit antigen specific T cell reactivity. This strategy may be useful for the induction of apoptosis in allopeptide reactive T cells driving chronic rejection.
5- Salvat R, Moise L, Bailey-Kellogg C, Griswold K E. J Vis Exp. (85):51308 (2014)
Biochemical assays with recombinant human MHC II molecules can provide rapid, quantitative insights into immunogenic epitope identification, deletion, or design(1,2). Here, a peptide-MHC II binding assay is scaled to 384-well format. The scaled down protocol reduces reagent costs by 75% and is higher throughput than previously described 96-well protocols(1,3-5). Specifically, the experimental design permits robust and reproducible analysis of up to 15 peptides against one MHC II allele per 384-well ELISA plate. Using a single liquid handling robot, this method allows one researcher to analyze approximately ninety test peptides in triplicate over a range of eight concentrations and four MHC II allele types in less than 48 hr. Others working in the fields of protein deimmunization or vaccine design and development may find the protocol to be useful in facilitating their own work. In particular, the step-by-step instructions and the visual format of JoVE should allow other users to quickly and easily establish this methodology in their own labs.