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. 2020 Jan 10;10:2909. doi: 10.3389/fmicb.2019.02909

FIGURE 4.

FIGURE 4

Schematic representation of different strategies used for fluorescence-based detection of lectin and antibody (Ab) binding to bacterial carbohydrate and whole cell microarrays. The simplest setup involves incubation with a fluorescently labeled target (lower right side). A common strategy is the use of biotinylated targets, which are next detected by incubation with fluorescently labeled streptavidin (upper part). The targets may carry other tags (e.g., His- or Fc-tags), and detection then has involved the use of biotinylated or unlabeled Abs, followed by incubation with streptavidin or with a biotinylated secondary Ab, as appropriate. Pre-complexing tagged targets with primary and secondary Abs has been exploited to reduce the number of incubation steps and/or to increase the sensitivity of detection. Alternatively, tagged targets have been detected by incubation with a fluorescent or unlabeled Ab, the latter followed by incubation with a fluorescent secondary Ab (lower part). Finally, the binding of unlabeled targets has been monitored using biotinylated or unlabeled primary Abs followed by fluorescently labeled streptavidin or secondary Abs. In all cases, the final step involves the scanning of fluorescence signals.