A statewide survey of questing ixodid ticks in mainland Florida was developed consistent with U.S. CDC standards to maximize the amount of epidemiologic and environmental data gathered. Survey sites were stratified by climatic zones and proportional to recognized land cover categories. A total of 560 transects on 41 sites within the state were sampled repeatedly by flagging between 2015 and 2018. Four tick species were collected; Amblyomma americanum, Amblyomma maculatum, Ixodes scapularis and Dermacentor variabilis. All species were more commonly found in northern and central regions of the state than in southern and western regions. Adult I. scapularis were active from autumn through spring and complementary to adult A. americanum and D. variabilis. Standardized survey methods help reduce sampling biases and better characterize risk from the species surveyed. However, differences in the attractiveness of collection methods for different tick species makes cross-species comparisons a continuing challenge.