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. Author manuscript; available in PMC: 2013 Oct 1.
Published in final edited form as: Mol Immunol. 2012 Jun 7;52(3-4):155–164. doi: 10.1016/j.molimm.2012.05.004

Figure 3. 2D methods for measuring in situ TCR-pMHC interactions.

Figure 3

(A) Single-molecule FRET. A TCR on the T cell membrane is labeled with a single chain antibody fragment conjugated with a FRET acceptor, and the peptide on a pMHC anchored to the lipid bilayer is labeled with a FRET donor. The TCR-pMHC interaction brings the donor and acceptor into close distance to trigger FRET. (B) Adhesion frequency assay. A T cell (right) is brought in and out of contact with a pMHC coated RBC (left) to estimate an adhesion frequency using micropipette manipulation. The presence or absence of RBC deformation signifies whether a TCR-pMHC bond is present on T cell retraction. (C) Thermal fluctuation assay. A T cell (right) is brought into close proximity to a pMHC coated bead attached to a RBC (left). Single bond association and dissociation events are detected by monitoring the thermal fluctuation amplitude of the bead that is either restrained by only the RBC or the combination of the RBC and the single TCR-pMHC bond.