Larval immersion tests (LIT) with commercial formulation of ivermectin were carried out with larvae of two field populations of Rhipicephalus (Boophilus) microplus from commercial dairy farms of the State of Sao Paulo, Brazil and a susceptible strain (Porto Alegre) to differentiate resistant-suspected and susceptible strains. One of the populations tested (Barra Alegre) showed a LC(50) value significantly higher than the susceptible strain and a resistance ratio (CI95%) of 3.78 (3.47-4.12), leading to suspect that this population shows traces of resistance to ivermectin. Population Sao Francisco, with no records of ivermectin injections on cattle, showed no difference on the ivermectin response in relation to Porto Alegre strain characterizing it as susceptible. Although LIT is not yet recommended by FAO to diagnose resistance to acaricides, this technique was successful on the differentiation of resistance-suspected population and a susceptible strain and can be used to detect populations of R. (B.) microplus resistant to ivermectin. This is the first report of R. (B.) microplus resistant to ivermectin detected by an in vitro bioassay.