Construction of a Heated Incubation Chamber around a Microscope Stage for Time-Lapse Imaging: Figure 1.

INTRODUCTIONIntravital imaging of embryogenesis has the potential to provide valuable information on cell proliferation, cell shape changes, and cell migratory behaviors. However, most embryo model systems require a temperature-controlled environment. Several expensive commercially available temperature control devices have emerged, including microscope stages surrounded by custom-fit Plexiglas boxes, heated plates for culture dishes, and objective warmers for water-immersion lenses, that strictly control temperature and, in some cases, help control local gas mixtures. This protocol describes an easy-to-assemble, cost-effective, custom-made cardboard box and incubator, adaptable to each user's specifications and microscope set-up. The cardboard box fits around the microscope, primarily the stage area, to assist in maintaining a prescribed temperature near the microscope stage. Warmed air, blown into the box enclosure from an incubator, circulates around the stage. The heated incubation box maintains a set temperature with minimal fluctuations and has been tested and utilized for studies of chick, mouse, and zebrafish embryogenesis.