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Immobilization of dye labeled proteins via biotin (protein)-streptavidin-biotin (surface) linkage to PEG-coated glass coverslip. Donor (Alexa Fluor 488) and acceptor (Alexa Fluor 594) dyes are labeled at the cysteine residues at positions 10 and 57 (Avi-GB1K10C/C57). Spacer and AviTag sequences with a biotin molecule were added at the N terminus of the protein.
