This chamber, top bars, and stage adapter are used to create a closed chamber for multiplex imaging, based on the chamber used in Radtke et al (2022). The chamber is placed on top of a large glass cover slip, with vacuum grease used to create a seal. To make a closed chamber, we have glued a small piece of glass to the top. There are two perfusion ports, one on each side, to allow for reagents to be added. The small reservoir is designed such that there can be continual suction without impacting the amount of liquid in the chamber. Four pins are required to secure the chamber and top bars to the stage adapter. Once on the adapter, the set can be placed on a Leica Thunder microscope.