title: Guiding synchrotron X-ray diffraction by multimodal video-rate protein crystal imaging.
abstract: Synchronous digitization, in which an optical sensor is probed synchronously with the firing of an ultrafast laser, was integrated into an optical imaging station for macromolecular crystal positioning prior to synchrotron X-ray diffraction. Using the synchronous digitization instrument, second-harmonic generation, two-photon-excited fluorescence and bright field by laser transmittance were all acquired simultaneously with perfect image registry at up to video-rate (15 frames s(-1)). A simple change in the incident wavelength enabled simultaneous imaging by two-photon-excited ultraviolet fluorescence, one-photon-excited visible fluorescence and laser transmittance. Development of an analytical model for the signal-to-noise enhancement afforded by synchronous digitization suggests a 15.6-fold improvement over previous photon-counting techniques. This improvement in turn allowed acquisition on nearly an order of magnitude more pixels than the preceding generation of instrumentation and reductions of well over an order of magnitude in image acquisition times. These improvements have allowed detection of protein crystals on the order of 1 µm in thickness under cryogenic conditions in the beamline. These capabilities are well suited to support serial crystallography of crystals approaching 1 µm or less in dimension.
authors.affiliation: Department of Chemistry, Purdue University, 560 Oval Drive, West Lafayette, IN 47906, USA.
authors.forename: Justin A
authors.initials: JA
authors.lastname: Newman
authors.affiliation: Department of Chemistry, Purdue University, 560 Oval Drive, West Lafayette, IN 47906, USA.
authors.forename: Shijie
authors.initials: S
authors.lastname: Zhang
authors.affiliation: Department of Chemistry, Purdue University, 560 Oval Drive, West Lafayette, IN 47906, USA.
authors.forename: Shane Z
authors.initials: SZ
authors.lastname: Sullivan
authors.affiliation: Department of Chemistry, Purdue University, 560 Oval Drive, West Lafayette, IN 47906, USA.
authors.forename: Ximeng Y
authors.initials: XY
authors.lastname: Dow
authors.affiliation: GM/CA@APS, X-Ray Science Division, Argonne National Laboratory, Argonne, IL 60439, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Becker
authors.affiliation: Department of Chemistry, Purdue University, 560 Oval Drive, West Lafayette, IN 47906, USA.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Sheedlo
authors.affiliation: GM/CA@APS, X-Ray Science Division, Argonne National Laboratory, Argonne, IL 60439, USA.
authors.forename: Sergey
authors.initials: S
authors.lastname: Stepanov
authors.affiliation: Department of Chemistry, Purdue University, 560 Oval Drive, West Lafayette, IN 47906, USA.
authors.forename: Mark S
authors.initials: MS
authors.lastname: Carlsen
authors.affiliation: Department of Chemistry, Purdue University, 560 Oval Drive, West Lafayette, IN 47906, USA.
authors.forename: R Michael
authors.initials: RM
authors.lastname: Everly
authors.affiliation: Department of Chemistry, Purdue University, 560 Oval Drive, West Lafayette, IN 47906, USA.
authors.forename: Chittaranjan
authors.initials: C
authors.lastname: Das
authors.affiliation: GM/CA@APS, X-Ray Science Division, Argonne National Laboratory, Argonne, IL 60439, USA.
authors.forename: Robert F
authors.initials: RF
authors.lastname: Fischetti
authors.affiliation: Department of Chemistry, Purdue University, 560 Oval Drive, West Lafayette, IN 47906, USA.
authors.forename: Garth J
authors.initials: GJ
authors.lastname: Simpson
mesh_terms: D018360:Crystallography, X-Ray; D007834:Lasers; D046911:Macromolecular Substances; D011506:Proteins; D017356:Synchrotrons; D014961:X-Ray Diffraction
chemical_list: D046911:Macromolecular Substances; D011506:Proteins
keywords: nonlinear optical microscopy; protein crystal; synchronous digitization