Summary: Green fluorescent protein (GFP) and its variants are believed to be the most powerful tool for protein visualization. However, due to their relatively large size and other drawbacks, many small molecular-based alternative methods have been developed over the years, of which bioorthogonal reactions are the most reliable and invaluable tools for protein labeling.1 Typically a bioorthogonal handle is first incorporated into a protein to enable site-specific modification. Then, various labels can be installed onto the protein via bioorthogonal reactions.
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