Further optimisation to improve crystal size and quality utilises a whole armoury of tips and tricks gained from many years of experience in X-ray structural determination from many different protein classes (e.g. kinases, phosphatases, proteases, receptors, Fab-antigen complexes and more). These include iterative variations of the precipitant formulation, seeding from poor quality crystals, varying the protein concentration and changing the temperature.
Protein crystals are then cryo-cooled in preparation for X-ray data collection at state-of-the-art synchrotron facilities. We have regular access at number of synchrotrons in Europe and also use specialist beamlines at these locations when appropriate. The output from data collection is progressed to structure solution via molecular replacement or de-novo phasing by Se-Met, SAD/MAD (when necessary). Multiple rounds of structure refinement follow using the industry standard CCP4 software package together with modelling and visualisation software such as Coot and PyMOL.