Binding Forces Of Single Alpha M Beta 2 Integrin-Fibrinogen Interactions On Living Cells

Single Cell Force Spectroscopy (SCFS) can be used to measure the maximum adhesion force between cells and a surface, but determining the nature of discrete interactions within SCFS data can be challenging. We present a method for quantifying specific ligand-receptor interactions on the single molecule level on living cells using atomic force microscopy (AFM) based SCFS experiments. SCFS data from HEK 293 cells expressing αMβ2 leukocyte integrin (HEK Mac-1) and wild-type HEK 293 (HEK WT) cells on surfaces coated with fibrinogen were analyzed to identify specific “rupture events.” High force load ruptures (u003e 0.2 pN/nm) imply a connection of the integrin with the underlying actin cortex of the cell, while low force load (u003c 0.2 pN/nm) ruptures result from the formation of a membrane tether. For highly adhesive fibrinogen surfaces, we found 41% of all rupture events to have a high force load for HEK Mac-1 cells compared to only 9% of rupture events having a high force load for HEK WT data of the same surface. The high force load events in the HEK Mac-1 data showed a median rupture force of 55 pN, whereas the median rupture force of the HEK WT high force load events was 29 pN. After adding monoclonal antibody directed against the αM subunit of the integrin, HEK Mac-1 cells showed similar rupture force values to that of the HEK WT. This analysis demonstrates the ability to quantify specific integrin-ligand interactions within SCFS data.