The present study investigated some in vitro properties for probiotic use of four strains of bifidobacteria isolated from faeces of healthy children (Bifidobacterium longum 5^1A, Bifidobacterium breve 110^1A, Bifidobacterium pseudolongum 119^1A and Bifidobacterium bifidum 162^2A). 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 5^1A 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 5^1A and B. pseudolongum 119^1A were able to produce inhibitory diffusible compounds against all pathogenic bacteria tested, but not against Candida albicans. B. longum 5^1A was sensitive to all the antimicrobials tested, except neomycin. The hydrophobic property of the cell wall was highest for B. bifidum 162^2A. Based on these parameters, B. longum 5^1A 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.