The present invention provides novel lyophilized rotavirus vaccine formulations and methods of their preparation. The formulations include vaccine stabilizers, resulting in a vaccine formulation with enhanced stability and minimal loss of potency. The rotavirus vaccine formulations comprise an advantageous ratio of a disaccharide (such as sucrose) to an amino acid (such as glycine). The lyophilization results in a virus formulation with 100% virus preservation and residual moisture from about 0.8% to 1.4%.