Detectors for imaging macro-molecules at atomic resolution

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

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

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

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

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

7. Detector requirements in structural biology Detective Quantum Efficiency : DQE = (Signalout/Noiseout)2/(Signalin/Noisein)2

8. Overview of detectors used in X-ray diffraction & electron microscopy

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 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 Fairchild CCD486 Courtesy Bruker-Nonius

13. X-rays Al p+ - E V d r i f t b i a s + n+ n++ Pixel detectors Courtesy MAR Research • 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 Christian Broennimann, PSI

14. MAR Research Solid State Direct Conversion detector

15. MAR Research Solid State Direct Conversion detector Dimensions: 430mm x 358mm Pixelsize: 140mm x 140 mm Number of pixels: 7.8 Mpixels Readout time: less that 1 s

16. MAR Research Solid State Direct Conversion detector

17. X-rays Al p+ - E V d r i f t b i a s + n+ n++ X-rays Sensor Chip Bump Bonds Ext /Comp Clock 15 bit RBI F SR 1 2 Clock counter RBO Gen Ext Clock 1.7fF Digital Block Analog Block Treshold correction - Global Comp + Tresh Bump Pad CS Amp Enable/ Reset Disable Cal Pixel Detectors: Principle Si pn -junction 3.6 eV to create 1 eh -pair Detector 0.2 mm Pixel Sensor 0.2 mm 0.3 mm Pixel electronics Pixel Read-out Chip Radiation hard

18. Paul Scherrer Institut PILATUS Detector with 3 Modules • Bank Data • Active Area: 238.7 x 35.3 mm2 • 157 x 1098 = 172386 pixels • 48 chips (radiation hard) • 2.38 mm gap between modules • Readout-time: 6 ms • Energy Range: Eg >4 keV • XY-addressing of each pixel • Threshold adjust of each pixel • Analog signal of each pixel Ch. Brönnimann

19. 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 Ch. Brönnimann

20. Future developments: digital holography? Single molecule diffraction Computa-tional phase retrieval Continuous or over-sampled diffraction pattern 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.