Abstract
We hypothesized that cancer cells in vivo might actively migrate towards micro vessels in tumor tissue along oxygen concentration gradients. To prove this hypothesis, an in vitro model of tissue oxygen gradients must be established. Here, we propose a very simple technique that may produce continuous gradients of oxygen concentration in cultured cells in conventional culture dishes. We devised the GCG (gap cover glass) by adhering two thin (150 µm) rectangular glass slips (9 x 18 mm) onto a 25 x 18 mm microscopy cover glass (the roof) in such a way that the glass slips are placed at interval of 2 mm on the roof cover glass. When the GCG is placed on cultured cells in a culture dish, supply of oxygen to the cell is limited only from the both open ends of the GCG. Furthermore, oxygen consumption by cells reduces oxygen concentrations under GCG as it enters from the open ends to the center. We tested whether gradients of oxygen are in fact established by using a thin film composed of oxygen sensitive (PtTFPP) and insensitive (N-(5- carboxypentyl)-4-piperidino-1,8-naphthalimide) fluorescent dyes.
