title: Nonlinear optical imaging of integral membrane protein crystals in lipidic mesophases.
abstract: Second-order nonlinear optical imaging of chiral crystals (SONICC) is explored for selective detection of integral membrane protein crystals grown in opaque and turbid environments. High turbidity is a hallmark of membrane protein crystallization due to the extensive use of detergent and/or lipids that often form various mesophases. Detection of crystals in such media by conventional optical methods (e.g., intrinsic UV fluorescence, birefringence, bright-field image analysis, etc.) is often complicated by optical scattering and by the small sizes of the crystals that routinely form. SONICC is shown to be well-suited for this application, by nature of its compatibility with imaging in scattering media and its high selectivity for protein crystals. Bright second harmonic generation (SHG) (up to 18 million counts/s) was observed from even relatively small crystals (5 mum) with a minimal background due to the surrounding lipid mesophase ( approximately 1 thousand counts/s). The low background nature of the resulting protein crystal images permitted the use of a relatively simple, particle counting analysis for preliminary scoring. Comparisons between a particle counting analysis of SONICC images and protocols based on the human expert analysis of conventional bright-field and birefringence images were performed.
authors.affiliation: Department of Chemistry, Purdue University, West Lafayette, Indiana 47907, USA.
authors.forename: David J
authors.initials: DJ
authors.lastname: Kissick
authors.forename: Ellen J
authors.initials: EJ
authors.lastname: Gualtieri
authors.forename: Garth J
authors.initials: GJ
authors.lastname: Simpson
authors.forename: Vadim
authors.initials: V
authors.lastname: Cherezov
mesh_terms: D003460:Crystallization; D005456:Fluorescent Dyes / Q000737:chemistry; D006801:Humans; D008055:Lipids / Q000737:chemistry*; D008565:Membrane Proteins / Q000737:chemistry*; D055620:Optical Phenomena
chemical_list: D005456:Fluorescent Dyes; D008055:Lipids; D008565:Membrane Proteins