FOXOs are highly dynamic transcription factors consisting of one conserved DNA-binding domain (forkhead domain) as well as intrinsically disordered regions (IDRs) at the N- and C-termini. These IDRs are essential and regulate transcriptional activity of FOXOs by serving as interaction platform for cofactors. Furthermore, the IDRs are involved in intra- and intermolecular homeotypic and heterotypic interactions between FOXOs and in turn mediate FOXO auto-inhibition and condensate formation. Here, we describe generalizable methods to study the structure and dynamics of FOXO proteins in vitro using a combination of biophysical techniques, in particular nuclear magnetic resonance spectroscopy. These methods can serve as a blueprint for the investigation of other IDR-containing transcription factors.