In the present study, an attempt was made to formulate and evaluate mucoadhesive bilayer buccal tablets of amphotericin B (antifungal agent), in order to overcome bioavailability problems, to reduce dose dependent side effects and frequency of administration. Mucoadhesive bilayer buccal tablets of amphotericin B were prepared by direct compression method by using combination of polymers HPMC and ethyl cellulose as backing layer. Five formulations were developed with varying concentrations of polymer. The designed tablets were evaluated for various parameters like hardness, friability, drug content, swelling index, mucoadhesion strength, in vitro drug release. Among five formulations, the formulation containing containing HPMC (100 mg), ethyl cellulose (50 mg), magnesium sterate (2 mg), sodium lauryl sulphate (4 mg), micro crystalline cellulose (44 mg) was selected as a best formulation. This formulation exhibited an in vitro drug release of 96.19% in 8 hrs along with satisfactory mucoadhesion strength (6.24 g). The prepared mucoadhesive bilayer buccal tablets of amphotericin B were able to stay in the buccal cavity for a longer period of time, which indicates a potential use of mucoadhesive bilayer tablets of amphotericin B for treating fungal infection.