Rhipicephalus microplus is among the critical tick parasite of cattle farming globally. However, the massive use of ivermectin last decades induced the generation of the tick-resistant population. Unfortunately, despite extensive research the mode of action of ivermectin remains unclear. Due to the limited molecular information related to the resistance of R. microplus to ivermectin, we carried out a midgut comparative proteomics study between susceptible and resistant tick. Our tissues specificity approach also included tandem mass tags (TMT) couple to synchronous precursor selection (SPS)-MS3. Our proteomics scrutiny exhibits key biological process associated with blood digestion and anticoagulation in midgut. In addition, we could observe the significant shutdown of the translation mechanism in susceptible thinks. At the same time, resistant thick exhibited the over-accumulation of cytochrome P450, glutathione-S-transferase, and ABC transporters. A similar trend was observed in proteins like glutathione peroxides glutaredoxin, sulfiredoxin, and disulfide-isomerase, which suggest a possible activation of the oxidative metabolism as a potential strategy to cope with ivermectin treatment.