Coffea arabica is one of the most important crops worldwide. In vitro culture is an alternative for achieving Coffea regeneration, propagation, conservation, genetic improvement, and genome editing. The aim of this work is to identify proteins involved in auxin homeostasis by isobaric tandem mass tag (TMT) and the synchronous precursor selection (SPS)-based MS3 technology on the Orbitrap Fusion™ Tribrid mass spectrometer™ in three types of biological material corre-sponding to C. arabica: plantlet leaves, calli, and suspension culture. Proteins included in the β-oxidation of indole butyric acid, and in the signaling, transport, and conjugation of in-dole-3-acetic acid were identified, such as the indole butyric response (IBR), the auxin binding protein (ABP), the ATP-binding cassette transporters (ABC), the Gretchen-Hagen 3 proteins (GH3), and the indole-3-acetic-leucine resistant proteins (ILR). A more significant accumulation of pro-teins involved in auxin homeostasis was found in the suspension cultures vs the plantlet com-parison, followed by callus vs plantlet and suspension culture vs callus, suggesting greater par-ticipation of these proteins as cell differentiation increases.