Among the ectoparasites of cattle, Rhipicephalus (Boophilus) microplus (Canestrini) (Acari: Ixodidae) remains a major cause of economic losses to livestock. The chemical control with acaricides is still the most efficient method available to control ticks. The aims of this study were to diagnose resistance to amitraz in 16 tick populations from the States of São Paulo (14) and Paraná (2), using the larval immersion technique (LIT), and evaluate the effect of synergists [piperonyl butoxide (PBO), diethyl maleate (DEM), triphenyl phosphate (TPP)] on amitraz resistant and susceptible strains of cattle tick. Most of the evaluated populations (68.7 %) showed to be resistant to amitraz, with resistance ratios ranging from 2.14 to 132. The results suggest that the test procedure by LIT is sensitive and adequate for detection and monitoring of amitraz resistance in cattle tick. No synergistic effect was observed for the synergists PBO, DEM and TPP, on the amitraz resistant (Poa) strain of cattle tick, indicating that increased detoxification metabolism was not involved in this resistance.