Mike Barnkob:Protocols/Immunology/Making tetramer

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Making Tetramers

Protocol for conjugating monomers to streptavidin-APC.

Reagents and equipment

1. Monomers
2. Straptavidin-APC (or other streptavidin-flurochrome conjugate)

Protocol

1. Calculate: Volume of Streptavidin = volume of monomer x 1.2
   Note: For 50μL monomer = 60 μL streptavidin.
2. Adjust concentration of both monomer and streptavidin to 1 mg / mL.
3. Add 1/3 of streptavidin volume to monomer and leave at 4°C O/N or 2 hours. Keep in the dark.
   Note: Carefully add the streptavidin. Do not pipet much up and down, and do not vortex.
4. Add 1/10 of streptavidin volume to monomer every 20 minutes. Keep at 4°C and in the dark.

References

• Tricks with tetramers: how to get the most from multimeric peptide–MHC
• Tetramerization
• Conjugation of monoclonal antibodies