A 2D Covalently Bound Continuous Protein Gradient Assay

Abstract

ABSTRACTTraditional methods of quantifying cell movement in response to a chemotactic factors provide either a binary count of cell migration in response to a known concentration of the factor of interest in solution, as in Boyden chamber assays, or a method of tracking cells to determine velocities across a solubilized protein gradient where exact concentrations vary over time and are difficult to define, as in the Ibidi chemotaxis gradient assay. Using a silane self-assembling monolayer (SAM)-based procedure pioneered by V Hlady and associates, we have developed an assay capable of covalently binding a wide variety of proteins to an optically transparent surface in a 2D pattern via amine linkages. The pattern was then verified by contact angle and Raman and X-ray photoelectron spectroscopy. This new assay provides greater control of protein concentration and gradient intensity than when using only solubilized proteins.