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T cells use sophisticated shape dynamics (morphodynamics) to migrate towardsand neutralise infected and cancerous cells. However, there is limitedquantitative understanding of the migration process in 3D extracellularmatrices (ECMs) and across timescales. Here, we leveraged recent advances inlattice light-sheet microscopy to quantitatively explore the 3D morphodynamicsof migrating T cells at high spatiotemporal resolution. We first developed anew shape descriptor based on spherical harmonics, incorporating keypolarisation information of the uropod. We found that the shape space of Tcells is low-dimensional. At the behavioural level, run-and-stop migrationmodes emerge at ~150 s, and we mapped the morphodynamic composition of eachmode using multiscale wavelet analysis, finding 'stereotyped' motifs. Focusingon the run mode, we found morphodynamics oscillating periodically (every ~100s) that can be broken down into a biphasic process: front-widening withretraction of the uropod, followed by a rearward surface motion and forwardextension, where intercalation with the ECM in both of these steps likelyfacilitates forward motion. Further application of these methods may enable thecomparison of T cell migration across different conditions (e.g.differentiation, activation, tissues, and drug treatments), and improve theprecision of immunotherapeutic development.

T cell morphodynamics reveal periodic shape oscillations in 3D migration