It has been proposed that the virulence of nosocomial Staphylococcus infections associated with indwelling medical devices is related to the ability of the bacterium to colonise these materials by forming a biofilm composed of multilayered cell clusters embedded in a slime matrix. However, the pathogenic role of exopolysaccharide biofilms is not fully understood. A new method was sought for differentiating the structure of slime from two closely related bacterial strains, Staphylococcus aureus and Staphylococcus epidermidis. Using PCR it was confirmed that these strains were positive for the icaA and icaD genes and the complete ica operon (2.7 kb). Monosaccharide analysis by thin-layer chromatography revealed an identical profile for both strains, with xylose and glucose present among the four visible bands. Using Fourier-transformed infrared spectroscopy and hierarchical cluster analysis, three of four S. aureus samples (75%), and four of five S. epidermidis samples were grouped according to species. A novel FTIR approach in classifying slime produced by S. aureus and S. epidermidis is reported.