The NIH Tetramer Facility, established in 1999, provides custom synthesis and distribution of soluble MHC-peptide tetramer reagents that can be used to stain antigen-specific T cells. MHC /peptide tetramer reagents are produced at the NIH Tetramer Facility contract site located at Emory University. MHC Allele protein production, folding and quality control are free to approved investigators (see Request Prioritization). Requestors incur the cost of peptide production (where required), which they supply to the facility (see General Guidelines) and shipping. All reagents are quality controlled (see General Guidelines) and are labeled with commercially manufactured streptavidin-allophycocyanin or streptavidin-phycoerythrin fluorophores. The NIH Tetramer Facility will not provide tetramers sold by Beckman Coulter, Inc.- Immunomics Operations (BCI). For a listing of the commercially-available tetramers, please refer to the Request Form home page:
http://research.yerkes.emory.edu/tetramer_core/request/requestform.html
Class I tetramers: A full list of available alleles can be found at:
http://research.yerkes.emory.edu/tetramer_core/allele/alleles.html Approved requestors must provide 10 (ten) milligrams of HPLC purified peptide (80% or greater purity) for tetramer synthesis. All exceed peptide will be stored for use only by the investigator supplying the peptide.
In addition to Class I MHC tetramer production, the NIH Tetramer Facility also provides: (1) Human CD1a, CD1b, and CD1c tetramers; and human and mouse CD1d tetramers; (2) Custom and pre-made class II MHC reagents for four distinct HLA-DR alleles; and (3) Tetramers with an expanded range of fluorophores.
CD1d reagents: Human and mouse CD1d molecules as tetramer reagents loaded with PBS-57, an analogue of alpha-GalCer ligand. PBS-57 is a chemical compound that was recently developed by Dr. Paul B. Savage, Brigham Young University (manuscript in preparation). Lymphocyte staining data is available at: http://research.yerkes.emory.edu/tetramer_core/CD1d_Tetramers.html
Two alpha-galactosylceramide analogues are now available for distribution: OCH and alpha-C-galactosylceramide. These CD1d ligands have previously been shown to stimulate natural killer T cells. All of these reagents are to be used for non-commercial research purposes only and may not be used in humans or for clinical diagnosis or treatment decisions.
OCH: Through a collaboration with Dr. Paul Savage at Brigham Young University, the NIH Tetramer Facility provides purified OCH (an alpha-galactosylceramide analogue with a truncated side chain). This compound stimulates Th2-biased cytokine production in NK T cells and has been demonstrated to delay the onset of experimental autoimmune encephalomyelitis in an animal model (Miyamoto et al., Nature, 2001). This ligand is available as a lyophilized powder derived from a Tween/sucrose/histidine-containing solution that can be reconstituted in water and is suitable for in vitro cell stimulations and in vivo animal studies (final concentration = 0.2 mg/mL). DMSO stock solutions of OCH without Tween/sucrose/histidine are also available. OCH-loaded CD1d tetramers can be made, but PBS-57-loaded reagents are recommended because they have been shown to stain NKT cells more brightly (Paul Savage, personal communication).
alpha-C-galactosylceramide: Through a collaboration with Dr. Richard Franck at Hunter College of CUNY, the NIH Tetramer Facility is offering purified alpha-C-galactosylceramide (a synthetic analogue of alpha-galactosylceramide). This C-glycoside is a potent stimulator of natural killer T cells and has been shown to protect animals against certain infections and cancers (Schmieg et al., J Exp Med, 2003). This compound is available in 1 mg aliquots for in vitro NK T cell activation. This reagent is to be used for non-commercial research purposes only and may not be used in humans or for clinical diagnosis. We are not able to provide alpha-C-galactosylceramide-loaded CD1d tetramers due to the instability of these reagents.
MHC class II Tetramers: The Tetramer Core Facility currently offers a variety of pre-made and custom human and mouse class II tetramers, and one non-human primate reagent. For a full list of available reagents, please see: http://research.yerkes.emory.edu/tetramer_core/availbMHCclassII.html
Pre-assembled MHC Class II tetramers: Fifteen pre-made class II reagents are available, including 11 mouse, 2 human, and 2 chimp reagent. All of these reagents are made with the peptide tethered to the MHC class II beta chain. A complete list of available pre-made class II tetramers can be found on the request form: http://research.yerkes.emory.edu/tetramer_core/request/ClassIIrequestform.html
Custom Class II MHC Tetramers: The Custom Class II Tetramer order form is located at:
http://research.yerkes.emory.edu/tetramer_core/request/customClassIIrequest.html ; and two main classes of reagents are available
(1) Human Class II tetramers made through CLIP peptide exchange: 4 HLA-DR alleles, provided through a collaboration with Dr. Kai Wucherpfennig (Day et al. (2003) J. Clin. Invest. 442: 831-42) at the Dana Farber Cancer Institute:
DRB1*0101/DRA1*0101;
DRB5*0101/DRA1*0101;
DRB1*1501/DRA1*0101;
DRB1*0401/DRA1*0101.
The requestor provides the peptide (0.5 mg) of interest and pays shipping costs; tetramer production costs are covered by the Tetramer Facility.
(2) Human and mouse MHC class II tetramers made with the peptide tethered to the MHC beta chain: I-A(b); I-A(d); I-A(g7); DRB1*0301/DRA1*0101. Approved investigators do not supply peptide. Reagent production can take several months after approval, since a new cell line expressing the peptide of interest must be made for each unique request.
Tetramer Facility staff are currently subcloning a number of class II genes for future tetramer production. In order to help the NIH Tetramer Facility prioritize these efforts, please complete the class II epitope survey and tell us which alleles are most valuable for your research. The survey is available at:
http://research.yerkes.emory.edu/tetramer_core/classII/ClassIIdatabase.html
Expanded Range of Fluorophores: In addition to the standard PE- and APC-labeled tetramers, you can now order class I tetramers with the following labels: Pacific Blue; Fluorescein; Alexa 488 (similar in fluorescence emission to fluorescein); Alexa 647 (similar in fluorescence emission to APC); and Alexa 680
