Biomembrane Force Probe to Quantitate Receptor-Ligand Interactions
Transcript
The interaction between a T-cell receptor, TCR, and an antigenic peptide presented by the MHC, or major histocompatibility complex, on an antigen-presenting cell, APC, causes the T-cell to exert a mechanical force on the APC. This phenomenon is necessary for T-cell activation.
To quantify receptor-ligand interactions using a biomembrane force probe, BFP, begin with a slide-based custom cell chamber. Place the cell chamber under a microscope.
Inject suspensions of biotinylated swollen RBCs, probe glass beads coated with peptide-bound MHC-streptavidin-biotin complexes, and T-cells into the chamber. Position the probe, target, and helper buffer-filled micropipettes, around the chamber.
Aspirate a biotinylated RBC with the probe pipette, positioning it in a blank area. Aspirate a T-cell with the target micropipette, positioning it on the RBC's right. Aspirate a probe bead with the helper micropipette, and attach it to the RBC's apex. The bead's streptavidin binds to the RBC's biotin and forms a force probe.
Align the T-cell with the force probe to allow the TCR to interact with its ligand ― the MHC-bound peptide ― and the T-cell to exert force on the RBC. Retract the T-cell to break the TCR-peptide bond, which deforms the RBC and displaces the bead.
Measure the force needed to break the bond ― a stronger bond needs a higher force and more time to break, indicating a longer lifetime.
