The paper describes the criteria, based on the multiple frequency analysis, used in the Italian expeditions to the Ross Sea to discriminate Euphausia superba (krill) aggregations from Euphausia crystallorophias ones and from other scatters. They are based on the experimental data acquired in the expeditions 1989/90, 1997/98 and 1999/2000. These criteria are presented, discussed, compared with literature and applied to estimate krill biomass in the area of Ross Sea (22000 nm2), investigated from 19 to 26 December 1997. It results that krill biomass estimated by the criteria based on multiple frequency method is 8.87% lower than krill biomass estimated using conventional methods (net samplings and/or echogram analysis).