While thick stress fibers are preserved well by most fixatives, many fine components of actin cytoskeleton are sensitive to physical and chemical perturbations and are damaged, destroyed or not completely preserved by commonly used fixation procedures. relies either on combination of electron and fluorescence microscopy, or on super-resolution fluorescence microscopy. This review focuses on approaches and techniques used to specifically reveal synaptic organelles and protein complexes, such as cytoskeletal assemblies. We place the strongest emphasis on methods detecting the targets of interest by affinity binding, and we discuss the advantages and limitations of each method. (McCann and Lichtman, 2008). Besides acetylcholine receptors, alpha-bungarotoxin was also used for studying AMPA receptor and GABA receptors localization and trafficking in neurons (Sekine-Aizawa and Huganir, 2004; Wilkins et al., 2008; Brady et al., 2014). In the latter studies, the alpha-bungarotoxin-binding site was fused to the proteins of interest, to enable the toxin to recognize receptors it usually does not bind to. This makes the bungarotoxin-binding site an affinity tag which, in principle, can be used for specific visualization of any membrane protein as long as adding this tag does not change the receptor targeting and trafficking. Another group of neurotoxins that started to be used for postsynaptic receptor visualization more recently is conotoxins C small peptides of 10C30 DNQX amino acids found in the venom of the snails. Various types of conotoxins were identified, each having a high affinity to a different target protein, including nicotinic acetylcholine receptors (Nicke et al., 2004), voltage-gated sodium channels (Leipold et al., 2005), potassium channels (Shon et al., 1998), and calcium channels (Nielsen et al., 2000). These small peptides can be conjugated chemically to fluorescent dyes and used as small probes to label respective proteins (Vishwanath and McIntosh, 2006). Very similar in structure, a component of deathstalker scorpion venom chlorotoxin has high affinity for chloride channels (DeBin et al., 1993). Many other scorpion venom components are used to study channels and receptors and can also be produced as recombinant fluorescent proteins to be used in microscopy (Kuzmenkov et al., 2016). While these toxins provide very high affinity and specificity, working in nano- and picomolar concentrations and being able to distinguish between very similar classes of receptors, their small size often makes it difficult and expensive to label them with fluorescent reporters, thus limiting their use. Labeling Proteins With Small Affinity Tags When no specific binder exists for a target protein, and fusion with a fluorescent protein has to be avoided, small peptide tags can be used to specifically visualize such proteins. They are short sequences of several amino acids that can be fused to any protein of interest and then targeted by a strong specific binder. The FLAG-, HA-, and myc-tags (Evan et al., 1985; Hopp et al., 1988; Wilson et al., 2005) are ones of the most commonly used affinity tags in imaging. Due to their small size (1.1 kDa) they are not expected to drastically affect the proteins traffic or function, and can be visualized by any imaging method following a staining with antibodies labeled with a suitable fluorophore. To increase the brightness of labeling, several copies of one tag can be fused to a protein, resulting in several antibodies binding to one target. When expressed on the extracellular domains of the plasma membrane proteins, these tags can be used for live cell imaging and tracking, as in the case of discussed above bungarotoxin-binding sites. However, the bivalency of the antibodies might introduce artifacts caused by protein clusters formation. The large size of the antibodies also restricts their ability to penetrate DNQX into confined and crowded environments, and can affect protein trafficking when applied to live cells. To solve these issues, smaller monovalent IRAK3 binders can be used. One possible alternative is monomeric streptavidin (Chamma et al., 2016a). To be recognized by streptavidin, the target protein must be fused to a 15 amino DNQX acid biotinylation substrate peptide (AP-tag). When biotin and biotin ligase are added to the cellular medium, the AP-tag is biotinylated and can be specifically bound by streptavidin. In addition to having an advantage of not cross-linking the target proteins, monomeric streptavidin is also substantially.