1. Detectors for imaging macro-molecules at atomic resolution JP Abrahams, Biophysical Structural Chemistry, Leiden University With thanks to: Jules Hendrix, MAR Research, Hamburg Christian Broennimann, PSI, Villingen Diederik Ellerbroek, Bruker-Nonius, Delft

2. Biophysical Structural Chemistry Genome Proteome information substance gene organism molecule Cell Central question: how do molecules interact to create life?

3. Research in life science ORGANISM physiology CELL cell biology MOLECULES biochemistry / biophysics ATOMS structural biology Theory development Transfection: In vivo studies Extraction: in vitro studies array techniquesgenetics bio-informaticsproteomics Other techniques for identifying genes, proteins and/or ligands Genes, proteins metabolites

4. X-ray crystallography 1. Grow crystals 3. Solve phases and refine structure 2. Measure diffraction

5. Cryo-EM Generate a 3D reconstruction (Re-)align particles Identify particles EM images 3D model 20 nm 300 nm Courtesy: R Koning, J Plaisier, HK Koerten

6. focus diffraction image Optics of diffraction and imaging object detector lens diffraction Diffraction pattern

7. Detector requirements in structural biology Large sizeLow background+++/- Low point-spreadLow background+++ High DQE 1 Reduction of sample damage ++ High dynamic rangeLinear sampling of all intensities +++/- Fast readoutEfficient data collection++ Low costThere’s more to life than detectors ++ CharacteristicRequired for:diffractimage Detective Quantum Efficiency : DQE = (Signal out /Noise out ) 2 /(Signal in /Noise in ) 2

8. Overview of detectors used in X-ray diffraction & electron microscopy Chemical detection: photography obsolete state- of the art Multi-wireobsoletenever usedirrelevant Television camera: FAST obsoletenever usedirrelevant Image platestate of the art used sometimes not used CCDstate of the art up and coming Pixel detectorsnear future????? X-ray diffraction Electron diffraction Electron imaging

9. Image plate detectors Based on a system (storage phosphor) for medical applications Advantages: practically no intrinsic noise; large size high spatial resolution large dynamic range Disadvantages: long read-out time Manufacturers: MAR Research, Rigaku Technology is tried & trusted, no major future developments are foreseen MAR image plate detector

10. CCD detectors Photon detection of an X-ray phosphor by a CCD. Advantages: fast readout; low noise; reasonable spatial resolution Disadvantages: limited dynamic range; small size requires de-magnifying optics; reasonable spatial resolution; expensive Manufacturers: Bruker-Nonius, MAR Research, ADSC Technology is recent and still developing Courtesy Bruker-Nonius

11. CCD detectors – new developments: Lens-based de-magnification Courtesy Bruker-Nonius MAR-research

12. CCD detectors – new developments: Next generation CCD’s Fairchild CCD486 JFET hybrid pre-amps: 2x faster, 2x lower noise Normal (buried channel) mode: 4x higher dynamic range Back illuminated CCD: 2-3X higher quantum efficiency Courtesy Bruker-Nonius

13. Pixel detectors Direct detection of electrons by pixel electrodes. Advantages: fast readout; low noise; high spatial resolution; high dynamic range Disadvantages: Very recent technology; first commercial products are anticipated for Autumn 2002 Manufacturers: MAR Research Technology is recent and still developing Courtesy MAR Research Courtesy Christian Broennimann, PSI

14. MAR Research Solid State Direct Conversion detector

15. Dimensions: 430mm x 358mm Pixelsize: 140  m x 140  m Number of pixels: 7.8 Mpixels Readout time: less that 1 s

16. MAR Research Solid State Direct Conversion detector

17. Pixel Detectors: Principle Pixel Sensor Pixel Read-out Chip 0.2 mm 0.3 mm Detector Radiation hard Si pn-junction 3.6eVto create 1eh-pair Pixel electronics Global Tresh Treshold correction CS Amp Comp Reset Bump Pad Enable/ Disable - + Cal 1.7fF Analog Block Digital Block 15 bit SR counter   Clock Gen RBI RBO Ext Clock Ext/Comp Clock

18. Ch. Brönnimann Paul Scherrer Institut PILATUS Detector with 3 Modules Bank Data Active Area: 238.7 x 35.3 mm 2 157 x 1098 = 172386 pixels 48 chips (radiation hard) 2.38 mm gap between modules Readout-time: 6 ms Energy Range: E  >4 keV XY-addressing of each pixel Threshold adjust of each pixel Analog signal of each pixel

19. Ch. Brönnimann Paul Scherrer Institut Diffraction pattern recorded with PILATUS Detector at Beamline 6s at the SLS Data Taking Lysozyme crystal 1 deg. Rotation (of a 45 deg data set) 2s exposure, E=12 keV Data taken at 7 detector positions Flatfield correction for each detector position

20. Future developments: digital holography? Single molecule diffraction Continuous or over-sampled diffraction pattern Computa- tional phase retrieval Courtesy Miao, Hodgson & Sayre, PNAS 98, p 6642

21. Summary & conclusions The ideal detector in structural biology has the following characteristics: For X-ray diffraction: Large, high-resolution, high dynamic range detectors with a fast readout. Detectors coming close to these specifications are available (CCD-based detectors) and more promising ones are around the corner (solid state direct conversion) For electron diffraction: Similar requirements as for X-ray diffraction. CCD detectors are already very good, but may be overtaken in future by solid state direct conversion detectors. For electron imaging: even higher resolution is required, but a high dynamic range is not as essential. It is not certain if direct conversion detectors will achieve a resolution that is sufficient; using large detectors will help, but this may require a re-think of the engineering of electron microscopes.