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The bio/muco-adhesive auxiliary agents are considered as a promising strategy for the administration of various challenging drugs including peptides, proteins and oligosaccharides therapeutics. The delivery of such therapeutics is hampered due to rapid degradation, restricted uptake, short residence time, poor solubility and limited half-life. The recent emergence of nanomedicine based on bio/mucoadhesive auxiliary agents is offering new avenues to overcome these drawbacks. Hence, it is important to understand the mechanism of nanoscale bio/mucoadhesion, protocols for investigating bio/mucoadhesive potential at nanoscale. This chapter is an endeavor to focus on the mechanism of bio adhesion and the bio/mucoadhesive auxiliary agents that are used in the design of the targeted oral, nasal, ocular, vaginal, and buccal nanomedicine with their properties that affect the bio/mucoadhesion.