Functionalized Bead Assay to Measure Three-dimensional Traction Forces during T-cell Activation

Area of Science:

• Immunology
• Cell Biology
• Biophysics

Background:

• T-cell receptor (TCR) and peptide-MHC interactions involve mechanical forces influencing antigen discrimination.
• Quantifying these forces during T-cell signaling is technically challenging.

Purpose of the Study:

• To develop a novel traction force microscopy platform for quantifying forces during T-cell activation.
• To investigate the role of T-cell microvilli in force exertion and cellular component reorganization.

Main Methods:

• Developed a traction force microscopy platform using functionalized marker beads.
• Immobilized T-cell activating antibodies on beads to measure hydrogel deformation.
• Utilized superresolution microscopy to confirm microvilli targeting and analyze actin organization.

Main Results:

• Quantified tangential and normal forces exerted by T-cell microvilli during activation.
• Observed reorganization of actin into a vortex-like structure around beads.
• Found enrichment of TCR and exclusion of CD45 at bead-microvilli contact sites.

Conclusions:

• The developed platform enables precise measurement of forces in T-cell signaling.
• Microvilli play a crucial role in force exertion and cellular component dynamics during T-cell activation.
• Force application leads to specific reorganization of key signaling molecules at the contact interface.