  The condensation of biomolecules into biomolecular condensates via
liquid-liquid phase separation (LLPS) is a ubiquitous mechanism that drives
cellular organization. To enable these functions, biomolecules have evolved to
drive LLPS and facilitate partitioning into biomolecular condensates.
Determining the molecular features of proteins that encode LLPS will provide
critical insights into a plethora of biological processes. Problematically,
probing biomolecular dense phases directly is often technologically difficult
or impossible. By capitalizing on the symmetry between the conformational
behavior of biomolecules in dilute solution and dense phases, it is possible to
infer details critical to phase separation by precise measurements of the
dilute phase thus circumventing complicated characterization of dense phases.
The symmetry between dilute and dense phases is found in the size and shape of
the conformational ensemble of a biomolecule - parameters that small-angle
x-ray scattering (SAXS) is ideally suited to probe. Recent technological
advances have made it possible to accurately characterize samples of
intrinsically disordered protein regions at low enough concentration to avoid
interference from intermolecular attraction, oligomerization or aggregation,
all of which were previously roadblocks to characterizing self-assembling
proteins. Herein, we describe the pitfalls inherent to measuring such samples,
the details required for circumventing these issues and analysis methods that
place the results of SAXS measurements into the theoretical framework of LLPS.
