The principle of fluorescent barcoding CLEM. (A) Schematic of MultiCLEM workflow: strains or cells in different experimental conditions are grown in parallel, and each population is stained with a unique combination of Con A conjugates (figure demonstrates three colors, but more can be used) to produce a combination (the barcode); strains are then mixed together and processed for EM. Samples are imaged with LM and EM at medium magnification (Mag), correlation is performed, cell identities are determined using the fluorescent barcode, and the high-resolution information is collected for each strain. (B) An example using three fluorophores to give seven well-discriminated combinations: pseudocolor composite image of a 100-nm-thick Lowicryl section of embedded cells labeled with Con A conjugated to Alexa 350, Alexa 488, and TRITC. (C) Examples of individual cells with all possible barcodes; each column corresponds to one fluorescent channel: R, TRITC; G, Alexa 488; B, Alexa 350. Scale bars: 10 µm in B, 2 µm in C.