Homemade confocal microscope environmental chamber for live cell imaging. (Left) Front view of the temperature- and humidity-controlled box that encloses the entire inverted microscope except for Hg arc and tungsten lamps. Orange letters indicate the separate pieces of the box: (H) Ultrasonic humidifier (Vicks); (hc) relative-humidity controller (RHCN-3A, Omega Engineering); (T) heater–air circulator (Air-Therm, World Precision Instruments). (Middle) The disassembled pieces of the box, which are made of 6-mm acrylic sheet, that fit snugly together and are locked into place with clasps. The front piece (F) has a rectangular aperture through which the eyepieces protrude (orange rectangle). The front and right half of the top (RT) are clear acrylic. The left and right sides (LS, RS), left half of the top (LT), and back panel (B) are opaque, black acrylic covered with reflective thermal insulating bubble wrap. The back panel and floor plate remain permanently mounted on the microscope. The remainder of the box disassembles for use at room temperature. (Upper right) Top view from the right side, with the top of the box removed. The condenser/lamp-housing post of the microscope tilts backward for access to the stage, pushing a swiveling panel that is set into the back panel of the box. (Lower right) Graph showing focal-plane shifts with different sample chambers. Blue and green lines represent shifts with a commercial sample chamber and objective-lens heater in auto and confocal modes. The red line represents the shifts with the homemade box. After an initial equilibration period, the focal plane with the homemade box is stable to within ∼0.2 µm.