TY - JOUR T1 - A Recipe for ORANGE-CAKE—This Time with Two Layers! JF - eneuro JO - eNeuro DO - 10.1523/ENEURO.0311-22.2022 VL - 9 IS - 4 SP - ENEURO.0311-22.2022 AU - Keely Duff Y1 - 2022/07/01 UR - http://www.eneuro.org/content/9/4/ENEURO.0311-22.2022.abstract N2 - As readers of eNeuro, we would agree that neurons are important cells—you are using them right now to read and understand this article [unless you are a sentient AI (artificial intelligence)–LaMDA (Language Model for Dialogue Applications); I would love to be friends!]. There are still a lot of unknowns about how neuronal proteins contribute to neuronal function. What exactly are the proteins in your neurons doing as you are reading this? We may not be able to determine that yet, but being able to visualize proteins in living neurons in culture is an important step toward that goal. The benefit of being able to “see” proteins is that you can determine where they go and how they move around in living cells. You can also get an idea of which proteins are interacting with each other.Fluorescent labeling of proteins has proven to be a useful tool for visualizing proteins in living cells. Historically, though, this method has had its drawbacks. In order to add a fluorescent tag to a protein, a gene encoding the tag and the protein of interest would be overexpressed in cells (typically done with a plasmid or virus). This can provide valuable information but can also result in atypical cellular function as it will alter normal protein … ER -