What are the limitations of molecular beacon probes?

One limitation is that once MB is degraded by an endogenous nucleus, the MB stem opens, generating false-positive signals. Additionally, the interaction of MB with intracellular proteins will interfere with the hairpin structure, interfering with the hairpin structure and resulting in non-specific signals. Another con is that they require a relatively large target sequence to ensure the complex is stable. Lastly, probe delivery is a drawback in the application of MBs in vivo as some delivery methods do not provide high efficiency and kinetics.