Acid and bile stability and intestinal mucosal adhesion properties are among the criteria used to select probiotic microbes. The quality control of probiotic cultures in foods traditionally has relied solely on tests to ensure that an adequate number of viable bacteria are present in the products throughout their shelf lives. Viability is an important factor, but not the only criterion for quality assurance. To be effective, probiotic strains must retain the functional health characteristics for which they were originally selected. Such characteristics include the ability to survive transit through the stomach and small intestine and to colonize the human gastrointestinal tract. In vitro test protocols can be readily adopted to examine the maintenance of a strain's ability to tolerate acidic conditions, survive and grow in the presence of bile, and metabolize selective substrates. Molecular techniques are also available to examine strain stability. Adhesion characterization may be an important quality-control method for assessing gut barrier effects. Adhesion has been related to shortening the duration of diarrhea, immunogenic effects, competitive exclusion, and other health effects. Adhesion properties should be carefully monitored, including adhesion to intestinal cells (eg, Caco-2) and human intestinal mucus. This article outlines the types of in vitro testing that can be used to ensure quality control of functional probiotic strains.