While recent advances in cryo-EM, coupled with single particle analysis, have the
potential to allow structure determination in a near-native state from vanishingly few individual
particles, this vision has yet to be realised in practise. Requirements for particle numbers that
currently far exceed the theoretical lower limits, challenges with the practicalities of achieving
high concentrations for difficult-to-produce samples, and inadequate sample-dependent imaging
conditions, all result in significant bottlenecks preventing routine structure determination using
cryo-EM. Therefore, considerable efforts are being made to circumvent these bottlenecks by
developing affinity purification of samples on-grid; at once obviating the need to produce large
amounts of protein, as well as more directly controlling the variable, and sample-dependent,
process of grid preparation.
In this proof-of-concept study, we demonstrate a further practical step towards this
paradigm, developing a 3D-printable flow-cell device to allow on-grid affinity purification from
raw inputs such as whole cell lysates, using graphene oxide-based affinity grids. Our flow-cell
device can be interfaced directly with routinely-used laboratory equipment such as liquid
chromatographs, or peristaltic pumps, fitted with standard chromatographic (1/16”) connectors,
and can be used to allow binding of samples to affinity grids in a controlled environment prior
to the extensive washing required to remove impurities. Furthermore, by designing a device
which can be 3D printed and coupled to routinely used laboratory equipment, we hope to
increase the accessibility of the techniques presented herein to researchers working towards
single-particle macromolecular structures.