The present study investigated some in vitro properties for probiotic use of four strains of bifidobacteria isolated from faeces of healthy children (Bifidobacterium longum 51A, Bifidobacterium breve 1101A, Bifidobacterium pseudolongum 1191A and Bifidobacterium bifidum 1622A). In vitro tests were carried out to compare growth rate, aerotolerance, antagonistic activity against pathogens, antimicrobial susceptibility profile and cell wall hydrophobicity. Mean doubling time of B. longum 51A was shorter compared to the other strains. All strains were aerotolerant up to 72 h of exposure to oxygen. In vitro antagonism showed that B. longum 51A and B. pseudolongum 1191A were able to produce inhibitory diffusible compounds against all pathogenic bacteria tested, but not against Candida albicans. B. longum 51A was sensitive to all the antimicrobials tested, except neomycin. The hydrophobic property of the cell wall was highest for B. bifidum 1622A. Based on these parameters, B. longum 51A showed the best potential for probiotic use among the tested strains, presenting the greatest sensitivity to antimicrobials, the best growth rate and the highest capacity to produce antagonistic substances against various pathogenic microorganisms.