RESUMO
We report a method for serial X-ray crystallography at X-ray free-electron lasers (XFELs), which allows for full use of the current 120-Hz repetition rate of the Linear Coherent Light Source (LCLS). Using a micropatterned silicon chip in combination with the high-speed Roadrunner goniometer for sample delivery, we were able to determine the crystal structures of the picornavirus bovine enterovirus 2 (BEV2) and the cytoplasmic polyhedrosis virus type 18 polyhedrin, with total data collection times of less than 14 and 10 min, respectively. Our method requires only micrograms of sample and should therefore broaden the applicability of serial femtosecond crystallography to challenging projects for which only limited sample amounts are available. By synchronizing the sample exchange to the XFEL repetition rate, our method allows for most efficient use of the limited beam time available at XFELs and should enable a substantial increase in sample throughput at these facilities.
Assuntos
Algoritmos , Cristalografia por Raios X/métodos , Aumento da Imagem/métodos , Interpretação de Imagem Assistida por Computador/métodos , Imageamento Tridimensional/métodos , Vírus/ultraestrutura , Reprodutibilidade dos Testes , Tamanho da Amostra , Sensibilidade e EspecificidadeRESUMO
Femtosecond vacuum ultraviolet (VUV) radiation provided by the free-electron laser FLASH was used for digital in-line holographic microscopy and applied to image particles, diatoms and critical point dried fibroblast cells. To realize the classical in-line Gabor geometry, a 1 microm pinhole was used as spatial filter to generate a divergent light cone with excellent pointing stability. At a fundamental wavelength of 8 nm test objects such as particles and diatoms were imaged at a spatial resolution of 620 nm. In order to demonstrate the applicability to biologically relevant systems, critical point dried rat embryonic fibroblast cells were for the first time imaged with free-electron laser radiation.
Assuntos
Elétrons , Holografia/métodos , Lasers , Raios Ultravioleta , Animais , Diatomáceas/citologia , Ratos , Dióxido de Silício , Fatores de Tempo , VácuoRESUMO
OBJECTIVE: To determine the feasibility of detecting the structural orientation in cartilage with Diffraction Enhanced X-Ray Imaging. DESIGN: Human tali and femoral head specimens were Diffraction Enhanced X-Ray Imaged (DEI) at the SYRMEP beamline at Elettra at various energy levels to detect the architectural arrangement of collagen within cartilage. DEI utilizes a monochromatic and highly collimated beam, with an analyzer crystal that selectively weights out photons according to the angle they have been deviated with respect to the original direction. This provides images of very high contrast, and with the rejection of X-ray scatter. RESULTS: DEI allowed the visualization of articular cartilage and a structural orientation, resembling arcades, within. CONCLUSION: Our diffraction enhanced images represent the first radiographic detection of the structural orientation in cartilage. Our data are in line with previous studies on the structural organization of joint cartilage. They confirm the model of a vaulting system of collagen fiber bundles interrupted by proteoglycan aggregates.