Method for In-Vitro Studies of Cellular Interactions at the Interface of Two Tissues

Abstract

We describe a simple and reliable experimental technique that enables one to create a high fidelity linear interface between two opposing cell layers. The method employs a custom designed lid that fits a standard 3cm cell culture dish. During cell plating, the dish is divided by a 200 micron thick separator that is part of the lid. The separator is covered in a thin layer of parafilm that forms a hermetic seal with the underlying coverslip and creates a temporary gap between the two cell plating environments. After cells attach, the custom lid is replaced with a standard lid and cells are allowed to grow under standard cell culture conditions. When expanding cell layers fill the gap, a linear interface is formed between the two opposing fields. Paracrinal factors released from an approaching cell front as well as direct physical and molecular interactions between two cell types affect intercellular orientation, individual cell morphology, and the degree of cells invasion into the opposing layer. The local interface appearance thus depends on a specific cell pair and may vary dramatically. We describe several types of such interfaces for different cell pairs, including cardiomyocytes, fibroblasts, melanocytes, endothelial cells and colon carcinoma cell lines. The method serves as a practical in vitro tool to study cell growth and invasion that occur on the interface of two neighboring tissues.