Preparation of Extracellular Matrices and Cell/Matrix Functional Units Produced by Cultured and Primary Fibroblasts

Abstract

Fibroblast‐derived matrices (FDMs) provide a biologically relevant in vitro alternative to conventional two‐dimensional (2D) substrates by enabling fibroblasts to assemble and organize their own extracellular matrix (ECM) environment, closely mimicking the physiological stromal microenvironment. This unit describes methods for generating interstitial matrices in which fibroblasts assemble their own multilayered extracellular scaffold, termed a fibroblast/ECM functional unit, that yields a cell‐free FDM anchored to the culture surface, following decellularization. The resulting FDM is a polymerized, native‐like scaffold that can be repopulated with cells, compressed, or solubilized for biomechanical and biochemical analysis. Unlike synthetic hydrogels or basement membrane extracts such as Matrigel, FDMs preserve native topological and mechanical features of the interstitial ECM.