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V. Motsch, M. Brameshuber, F. Baumgart, G. Schütz, E. Sevcsik:
"A micropatterning platform for quantifying interaction kinetics between the T cell receptor and an intracellular binding protein";
Scientific Reports, 9 (2019), 328801 - 328810.

A complete understanding of signaling processes at the plasma membrane depends on a
quantitative characterization of the interactions of the involved proteins. Fluorescence recovery
after photobleaching (FRAP) is a widely used and convenient technique to obtain kinetic parameters
on protein interactions in living cells. FRAP experiments to determine unbinding time constants for
proteins at the plasma membrane, however, are often hampered by non-specific contributions to the
fluorescence recovery signal. On the example of the interaction between the T cell receptor (TCR) and
the Syk kinase ZAP70, we present here an approach based on protein micropatterning that allows
the elimination of such non-specific contributions and considerably simplifies analysis of FRAP data.
Specifically, detection and reference areas are created within single cells, each being either enriched or
depleted in TCR, which permits the isolation of ZAP70-TCR binding in a straight-forward manner. We
demonstrate the applicability of our method by comparing it to a conventional FRAP approach.