A novel method for quantifying cell migration through tissue engineering scaffolds: evaluation of modified collagen matrices

Abstract

A novel method to quantify cell migration through potential tissue engineering 3-d scaffolds is described. The migration assay uses a dot-blotting apparatus into which the tissue engineering matrix is placed on top of a nitrocellulose membrane. This assay was used to evaluate human dermal fibroblast migration through four porcine collagen matrices with varying pore diameters and pitch lengths. Fibroblasts were placed on the matrix surface, at between 1 ×103–3 × 103 cells mm−2, and left for 18 h to allow migration. The nitrocellulose membrane was stained with haematoxylin, the membrane digitised and the pixel intensity of the stained cells quantified. We showed that for all matrix variants, migration was more effective with a higher initial seeding density. The application of varying initial cell densities resulted in the greatest extent of cell migration through the matrix variant with pores of 30 μm diameter and 400 μm pitch length (i.e. 10.3% migration at 1 ×103 cells mm−2). This method was coupled with confocal microscopy to evaluate the depth of cell migration within the matrix. At a depth of 20 μm cell numbers were similar to those on the matrix surface: at a depth of 100 μm only a few cells were observed.