Application of the dual-micropipet technique to the measurement of tumor cell locomotion

The objective of this work was to characterize tumor cell locomotion in response to chemotactic stimulation using a dual-micropipet assay. The assay involves two micropipets. An individual A2058 human melanoma cell was retained, without pressure gradient, in a pipet of approximately 14 μm i.d. A solution of type IV collagen, chosen as the chemotactic source, was placed in another pipet (~10 μm o.d.) with zero pressure at the pipet tip. The smaller pipet was then inserted into the larger one containing the melanoma cell. The initial chemoattractant concentration (C(o)) and the distance between the tip of the small pipet and the cell surface (δ) provided a gradient (C(o)/δ) for tumor cell locomotion toward stimulation. This novel assay provides a direct measure of cell movement: cyclic pseudopod protrusion (L(p)) and subsequent cell locomotion (L(o)). The influences of different adhesion substrates on cell locomotion were also studied. The peak length in L(p) precedes the highest locomotion velocity (dL(c)/dt) by an apparent lag time. C(o)/δ influences pseudopod protrusion frequency (f(p)) and dL(c)/dt, but not significantly on L(p). Substrate adhesions affect dL(c)/dt, but apparently not L(p) or f(p). In conclusion, pseudopod protrusion and substrate adhesion are two necessary but mutually independent factors in tumor cell locomotion, dL(c)/dt correlates with changes in C(o)/δ, which is in significant correlation with f(p) but not L(p).