Degassing in an automated RNA imaging instrumentation

The work in the Linnarsson Lab at Karolinska Institute, aims to make high resolution atlases of the mouse and human brain, where we know the location and type of every individual cell in the brain. This is done by imaging the RNA molecules that are present in each cell of the brain with a microscope. The RNA can be seen as an intermediate between the cells’ DNA and the proteins. The information of which specific RNA molecules are present in a cell tells us about the function of the cell. There are thousands of different RNA molecules that can be present in the cell which determine the identity of the cell, and in order to visualize such a high number of targets we perform many cycles of labeling and imaging on the same brain sample.

This takes an enormous amount of work and therefore we have automated the process, by building a fluidic machine that can dispense all the required liquids to the sample and start the imaging. We build three machines that are performing these experiments around the clock and there are many things that can go wrong in such a complex experiment. Air bubbles in the flow cell that holds the brain sample was a common reason why our experiments failed. By putting a bubble-trap and a degasser we can virtually catch and prevent air bubbles going into the flow cell. The degassing is especially important because the samples need to be incubated at different temperatures, and a liquid can quickly out-gas at higher temperatures, forming air bubbles that would have a detrimental effect on the system performance.

Apparatus: A modified analytical DEGASi^® Plus stand-alone degasser with different channel size to optimize degassing efficiency and system dead volume. The valves shown in the photo are IDEX MX Series II™ 10 Position/11 Port Selector Valves.

Author and system designer: Lars Borm | PhD student Linnarsson Lab, Karolinska Institute, Department of Medical Biochemistry and Biophysics, Unit of Molecular Neurobiology, Stockholm, Sweden, web link http://linnarssonlab.org/