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Electron microscopy in molecular cell biology I Electron optics and image formation Werner Kühlbrandt Max Planck Institute of Biophysics Donnerstag, 3. Juli 14 Objects of interest • • • • • • • • • • • • Galaxy! ! Solar system! Planet! ! Man! ! Fly! ! ! Cell ! ! Bacterium! ! Virus! ! Protein! ! Sugar molecule! Atom! ! Electron! ! Donnerstag, 3. Juli 14 ! ! ! ! ! ! ! ! ! ! ! ! 106 light years! light minutes! ! 20,000 km! ! 1.7 m! ! ! 3 mm! ! ! 50 µm! ! ! 1 µm! ! ! 100 nm! ! 10 nm! ! ! 1 nm! ! ! 0.1 nm = 1Å! ! 0.1 pm!! 1022 m 1010 m 107 m 100 m 10-3 m 10-5 m 10-6 m 10-7 m 10-8 m 10-9 m 10-10 m 10-13 m biology Objects of interest • • • • • • • • • • • • Galaxy! ! Solar system! Planet! ! Man! ! Fly! ! ! Cell ! ! Bacterium! ! Virus! ! Protein! ! Sugar molecule! Atom! ! Electron! ! Donnerstag, 3. Juli 14 ! ! ! ! ! ! ! ! ! ! ! ! 106 light years! light minutes! ! 20,000 km! ! 1.7 m! ! ! 3 mm! ! ! 50 µm! ! ! 1 µm! ! ! 100 nm! ! 10 nm! ! ! 1 nm! ! ! 0.1 nm = 1Å! ! 0.1 pm!! 1022 m 1010 m 107 m 100 m 10-3 m 10-5 m 10-6 m 10-7 m 10-8 m 10-9 m 10-10 m 10-13 m chemistry biology Objects of interest • • • • • • • • • • • • Galaxy! ! Solar system! Planet! ! Man! ! Fly! ! ! Cell ! ! Bacterium! ! Virus! ! Protein! ! Sugar molecule! Atom! ! Electron! ! Donnerstag, 3. Juli 14 ! ! ! ! ! ! ! ! ! ! ! ! 106 light years! light minutes! ! 20,000 km! ! 1.7 m! ! ! 3 mm! ! ! 50 µm! ! ! 1 µm! ! ! 100 nm! ! 10 nm! ! ! 1 nm! ! ! 0.1 nm = 1Å! ! 0.1 pm!! 1022 m 1010 m 107 m 100 m 10-3 m 10-5 m 10-6 m 10-7 m 10-8 m 10-9 m 10-10 m 10-13 m Which objects can we see? • • • • • • • • • • • • Galaxy! ! Solar system! Planet! ! Man! ! Fly! ! ! Cell ! ! Bacterium! ! Virus! ! Protein! ! Sugar molecule! Atom! ! Electron! ! Donnerstag, 3. Juli 14 ! ! ! ! ! ! ! ! ! ! ! ! 106 light years! light minutes! ! 20,000 km! ! 1.7 m! ! ! 3 mm! ! ! 50 µm! ! ! 1 µm! ! ! 100 nm! ! 10 nm! ! ! 1 nm! ! ! 0.1 nm = 1Å! ! 0.1 pm!! 1022 m 1010 m 107 m 100 m 10-3 m 10-5 m 10-6 m 10-7 m 10-8 m 10-9 m 10-10 m 10-13 m Resolution limit with visible light Which objects can we see? • • • • • • • • • • • • Galaxy! ! Solar system! Planet! ! Man! ! Fly! ! ! Cell ! ! Bacterium! ! Virus! ! Protein! ! Sugar molecule! Atom! ! Electron! ! Donnerstag, 3. Juli 14 ! ! ! ! ! ! ! ! ! ! ! ! 106 light years! light minutes! ! 20,000 km! ! 1.7 m! ! ! 3 mm! ! ! 50 µm! ! ! 1 µm! ! ! 100 nm! ! 10 nm! ! ! 1 nm! ! ! 0.1 nm = 1Å! ! 0.1 pm!! 1022 m 1010 m 107 m 100 m 10-3 m 10-5 m 10-6 m 10-7 m 10-8 m 10-9 m 10-10 m 10-13 m Structural methods in molecular cell biology light microscopy AFM NMR x-ray crystallography electron microscopy 0.1nm=1Å 1nm atoms 10nm proteins small molecules Donnerstag, 3. Juli 14 100nm 1µm 10µm bacteria viruses cells 100µm 1mm organisms 10mm Part 1: Electron optics Donnerstag, 3. Juli 14 Microscopy with electrons Donnerstag, 3. Juli 14 Microscopy with electrons Donnerstag, 3. Juli 14 Microscopy with electrons • Electrons are energy-rich elementary particles Donnerstag, 3. Juli 14 Microscopy with electrons • Electrons are energy-rich elementary particles • Electrons have much shorter wavelength than x-rays (~ 3 pm = 3 x 10-12 m for 100kV electrons) Donnerstag, 3. Juli 14 Microscopy with electrons • Electrons are energy-rich elementary particles • Electrons have much shorter wavelength than x-rays (~ 3 pm = 3 x 10-12 m for 100kV electrons) • Resolution not limited by wavelength! Donnerstag, 3. Juli 14 Microscopy with electrons • Electrons are energy-rich elementary particles • Electrons have much shorter wavelength than x-rays (~ 3 pm = 3 x 10-12 m for 100kV electrons) • Resolution not limited by wavelength! • but by comparatively poor quality of magnetic lenses and radiation damage Donnerstag, 3. Juli 14 Microscopy with electrons • Electrons are energy-rich elementary particles • Electrons have much shorter wavelength than x-rays (~ 3 pm = 3 x 10-12 m for 100kV electrons) • Resolution not limited by wavelength! • but by comparatively poor quality of magnetic lenses and radiation damage • High vacuum is essential - no live specimens! Donnerstag, 3. Juli 14 Electron microscopes 300 kV Donnerstag, 3. Juli 14 120 kV The transmission electron microscope camera Donnerstag, 3. Juli 14 The transmission electron microscope electron gun camera Donnerstag, 3. Juli 14 The transmission electron microscope electron gun condenser camera Donnerstag, 3. Juli 14 The transmission electron microscope electron gun condenser objective lens camera Donnerstag, 3. Juli 14 The transmission electron microscope electron gun condenser objective lens camera projector lenses Donnerstag, 3. Juli 14 The transmission electron microscope electron gun condenser objective lens camera projector lenses projection chamber Donnerstag, 3. Juli 14 The transmission electron microscope electron gun condenser objective lens camera projector lenses projection chamber Donnerstag, 3. Juli 14 vacuum pump The transmission electron microscope electron gun condenser objective lens camera projector lenses projection chamber viewing screen Donnerstag, 3. Juli 14 vacuum pump The transmission electron microscope electron gun condenser objective lens camera projector lenses projection chamber viewing screen camera Donnerstag, 3. Juli 14 vacuum pump The transmission electron microscope electron gun condenser objective lens specimen projector lenses projection chamber viewing screen camera Donnerstag, 3. Juli 14 camera vacuum pump The transmission electron microscope electron source specimen objective lens diffraction plane image plane focus plane Donnerstag, 3. Juli 14 The transmission electron microscope electron source specimen objective lens diffraction plane image plane focus plane Donnerstag, 3. Juli 14 Electron sources Thermionic emitters Tungsten filament: emits electrons at ~ 3000 °C (melts at 3380 °C) Large source size, poor coherence of electron beam LaB6 crystal: single crystal, emits electrons at ~ 2000 °C Electrons are emitted from crystal tip -> smaller source size, better coherence Energy spread ΔE ~ 1 - 2 eV, depending on temperature Field emitters Oriented tungsten single crystal, tip radius ~ 100 nm Small source size -> high coherence of electron beam, high brilliance Requires very high vacuum (10 -11 Torr) Electrons extracted from crystal by electric field applied to tip (extraction voltage) Schottky emitter: Zr plated, heated to ~ 1200 °C to assist electron emission and prevent contamination, energy spread ΔE ~0.5 eV Cold field emitter: not heated, very low energy spread, best temporal coherence, but contaminates easily (frequent ‘flashing’ with oxygen gas) Donnerstag, 3. Juli 14 Coherence tungsten filament field emission tip incoherent electron beam coherent electron beam temporal coherence: all waves have the same wavelength (they are monochromatic) spatial coherence: all waves are emitted from the same point source (as in a laser) Donnerstag, 3. Juli 14 Electrons: particles or waves? According to the dualism of elementary particles in quantum mechanics, electrons can be considered as particles or waves. Both models are valid and used in electron optics. Electrons can be regarded as particles to describe scattering. The wave model is more useful to describe diffraction, interference, phase contrast and image formation. Donnerstag, 3. Juli 14 Electron parameters − 31 Rest mass m 0 = 9.1091x10 Charge Kinetic energy Rest energy Q = −e = − 1.602x10 C − 19 E = eU ,1eV = 1.602x10 Nm E0 = m0c 2 = 511keV = 0.511MeV Velocity of li ght Planck’s constant c = 2.9979x10 ms −1 −34 h = 6.6256x10 Nms Non-relativistic (E << E0 ) Relativistic (E ~ E 0 ) Mass m = m0 m = m0 (1 − v 2 / c2 ) Energy E = eU = kg −19 8 1 2 m 0v 2 −1 / 2 mc 2 = m0 c2 + eU = E 0 + E m = m0 (1 + E / E0 ) 1/ 2 Velocity v = (2 E / m0 ) Momentum p = m 0v = ( 2m 0E ) v = c [1− 1/ 2 −1 / 2 h de Brogli e wavelength λ = = h (2 m0 E ) p 1 (1 + E / E0 ) 2 ] 1/ 2 p = mv = [ 2m 0 E (1 + E /2 E0 )] 1 2 1 /2 = ( 2 EE0 + E ) c λ = h [2m 0 E(1+ E / 2 E0 )] 2 = hc ( 2 EE0 + E ) −1 / 2 Donnerstag, 3. Juli 14 −1/2 1/2 Electrons in an electrostatic (Coulomb) field The force produced by an electrostatic field E on a particle of (negative) charge e- is F = -eE This means that the electron is accelerated towards the anode. The kinetic energy E [eV] of the electron after passing through a voltage U [V] is E = eU The wavelength of an electron moving at veolicty v (de Broglie wavelength) is λ = h/mv where h is Planck’s constant, m the rest mass of the electron. Both the mass and the velocity of the electron are energy dependent. Relativistic treatment is necessary for acceleration voltages > 80 kV. Electron beams are monochromatic except for a small thermal energy spread arising from the temperature of the electron emitter. Donnerstag, 3. Juli 14 de Broglie wavelength acceleration voltage 100 kV velocity 1.64 x 108 m/s wavelength 3.7 pm = 0.037 Å 200 kV 2.5 pm = 0.025 Å 300 kV 2.0 pm = 0.02 Å Donnerstag, 3. Juli 14 Resolution limit in crystallography Reciprocal space! Highest possible resolution: 1/d = 2/λ d = λ/2 θ = 90° Donnerstag, 3. Juli 14 Resolution limit Diffraction (Rayleigh) limit: ~ 0.5 λ • ~150 nm = 1,500 Å for visible light • 0.5 Å for x-rays • 0.01 Å = 1 pm for 300 kV electrons Donnerstag, 3. Juli 14 The transmission electron microscope electron source specimen objective lens diffraction plane image plane focus plane Donnerstag, 3. Juli 14 Electron-‐specimen interactions Incident beam of energy E hν E-ΔE E-ΔE Z=0 Z=t E E E -ΔE E elastic Donnerstag, 3. Juli 14 E -ΔE inelastic Elastic and inelastic scattering cross sections for electrons, x-rays and neutrons R. Henderson, Quart. Rev. Biophys. 1995 Donnerstag, 3. Juli 14 Electron scattering factors Large elastic cross section • ~106 higher than for x-‐rays high signal per scattering event … but even larger inelastic cross section • σinel/σel ≈ 18/Z high radiation damage, especially for light atoms, e.g. C: 3 electrons scattered inelastically per elastic event Good ratio of elastic/inelastic scattering • 1000 times better than for 1.5 Å x-‐rays Donnerstag, 3. Juli 14 carbon Considerations for biological specimens • Biological specimens consist mainly of light atoms (H, N, C, O), which suffer most from radiation damage Donnerstag, 3. Juli 14 Considerations for biological specimens • Biological specimens consist mainly of light atoms (H, N, C, O), which suffer most from radiation damage • This makes it essential to minimize the electron dose to about ~25 e/Å2 for single-‐particle cryoEM Donnerstag, 3. Juli 14 Considerations for biological specimens • Biological specimens consist mainly of light atoms (H, N, C, O), which suffer most from radiation damage • This makes it essential to minimize the electron dose to about ~25 e/Å2 for single-‐particle cryoEM • Low electron dose means low image contrast Donnerstag, 3. Juli 14 The transmission electron microscope electron source specimen objective lens diffraction plane image plane focus plane Donnerstag, 3. Juli 14 Electrons in a magnetic field Lorentz force on a charge e- moving with velocity v in a magnetic field B (vectors are bold): F = -e (v x B) The direction of F is perpendicular on v and B. v, B, and F form a right-handed system (right-hand rule, where v = index finger, B = middle finger and c = thumb) In a homogenous magnetic field, if v = 0 then F = 0 v is perpendicular to B: the electron is forced on a circle with a radius depending on the field strength |B| v is parallel to B: no change of direction Note that unlike the electric field, the magnetic field does not change the energy of the electron, i. e. the wavelength does not change! Donnerstag, 3. Juli 14 Electromagnetic field Passing a current through a coil of wire produces a strong magnetic field in the centre of the coil. The field strength depends on the number of windings and the current passing through the coil. Donnerstag, 3. Juli 14 Electromagnetic Lens Pole pieces of soft iron concentrate the magnetic field Donnerstag, 3. Juli 14 Electromagnetic Lens Pole pieces of soft iron concentrate the magnetic field Donnerstag, 3. Juli 14 Electromagnetic Lens Donnerstag, 3. Juli 14 Donnerstag, 3. Juli 14 Magnification steps 60x EM grid 2mm x 2mm Donnerstag, 3. Juli 14 Magnification steps 60x 600x EM grid grid square with holey carbon film 2mm x 2mm 20 µm x 20 µm Donnerstag, 3. Juli 14 Magnification steps 60x 6,000x 600x EM grid grid square with holey carbon film hole 2mm x 2mm 20 µm x 20 µm 2µm x 2µm Donnerstag, 3. Juli 14 Magnification steps 60x 6,000x 600x 60,000x EM grid grid square with holey carbon film hole FAS in vitreous buffer 2mm x 2mm 20 µm x 20 µm 2µm x 2µm 0.2 µm x 0.2 µm Donnerstag, 3. Juli 14 The transmission electron microscope electron source specimen objective lens diffraction plane image plane focus plane Donnerstag, 3. Juli 14 Electron diffraction d 2θ diffraction angle Bragg’s law: nλ = 2d sinθ Donnerstag, 3. Juli 14 Electron diffraction d 2θ diffraction angle Bragg’s law: nλ = 2d sinθ Donnerstag, 3. Juli 14 Interference constructive interference superposition 1st wave 2nd wave Donnerstag, 3. Juli 14 destructive interference Elastic scattering by atoms (Rayleigh scattering) n is an integer: scattered waves in phase, constructive interference n is not an integer: scattered waves out of phase, destructive interference Bragg’s law: nλ = 2d sinθ Donnerstag, 3. Juli 14 2D crystals of LHC-II Kühlbrandt, Nature 1984 Donnerstag, 3. Juli 14 Electron diffraction pattern: amplitudes, no phases 3.2 Å Donnerstag, 3. Juli 14 The image is generated by interference of the direct electron beam with the diffracted beams Donnerstag, 3. Juli 14 Part 2: Image formation Donnerstag, 3. Juli 14 The transmission electron microscope electron source specimen objective lens diffraction plane image plane focus plane Donnerstag, 3. Juli 14 Electron-‐specimen interactions Incident beam of energy E hν E-ΔE E-ΔE Z=0 Z=t E E E -ΔE E elastic Donnerstag, 3. Juli 14 E -ΔE inelastic Contrast mechanisms electron wave Image no interaction modified amplitude, unchanged phase amplitude object amplitude contrast Donnerstag, 3. Juli 14 phase contrast phase object shorter wavelength within object modified phase modified phase, unchanged amplitude Origin of the phase shift The potential V(x,y,z) experienced by the scattered electron upon passing through a sample of thickness t causes a phase shift t Vt (x, y) = ψincident ∫ V (x, y,z)dz 0 The vacuum wavelength λ of the electron changes to λ` within the sample: V(x,y,z) ψexit Donnerstag, 3. Juli 14 € € € h λ= 2meE € λ" = h 2me(E + V (x, y,z)) When passing through a sample of thickness dz, the electron experiences a phase shift of: € dz dz (how many times does λ fit into the thickness dz?) dϕ = 2π ( − ) (by how many degrees out of 2π = 360° does λ$ λ this change the phase φ?) π dϕ = V (x, y,z)dz = σV (x, y,z)dz λE The weak phase approximation As we have seen, the phase shift is proportional to the projected potential of the specimen: ϕ = σ ∫ V (x, y,z)dz = σVt (x, y) The potential V of the specimen shift modifies the phase of the incident plane wave ψincident to: € ψincident Ψexit = Ψincident e −iϕ = Ψincident e −iσVt For Vt<<1 and thus φ<<2π as for thin biological specimens: V(x,y,z) € i =e i π2 = cos( π2 ) + isin( π2 ) Ψexit = Ψincident e −iϕ ≈ Ψincident (1 − iϕ) = Ψincident − iΨincidentϕ = Ψincident − iΨscattered € ψexit € àFor thin biological specimens there is a linear relation between the transmitted exit wave function and the projected potential of the sample: Ψexit ≈ Ψ !incident "# (1 − iϕ) = 1 − iϕ = 1 − iσVt ≡1(background ) Donnerstag, 3. Juli 14 Taylor series Any function can be approximated by a finite number of polynomial terms ∞ à ∑ n =0 f (n ) (a) (x − a) n n! à à e x ≈ 1+ x + à e −iϕ ≈ 1 − iϕ + € € € x3 x5 x7 + − + O(x n ) 3! 5! 7! sin(x) ≈ x − € Donnerstag, 3. Juli 14 x2 x3 + + O(x n ) 2! 3! ϕ2 ϕ3 − i + O(ϕ n ) 2! 3! ≈ 1 − iϕ if φ << 1 ! taken from Wikipedia... Taylor series Any function can be approximated by a finite number of polynomial terms ∞ à ∑ n =0 f (n ) (a) (x − a) n n! à à e x ≈ 1+ x + à e −iϕ ≈ 1 − iϕ + € € € x3 x5 x7 + − + O(x n ) 3! 5! 7! sin(x) ≈ x − € Donnerstag, 3. Juli 14 x2 x3 + + O(x n ) 2! 3! ϕ2 ϕ3 − i + O(ϕ n ) 2! 3! ≈ 1 − iϕ if φ << 1 ! taken from Wikipedia... Phase contrast exit wave = scattered wave + unscattered wave imaginary Scattered wave is 90° out of phase with unscattered wave. Ψexit = ψunscattered + iψscattered exit wave real Vector notation. Vector length shows amplitude (not drawn to scale) Donnerstag, 3. Juli 14 Cosine wave notation (amplitudes not drawn to scale) Phase contrast exit wave = scattered wave + unscattered wave A perfect lens in focus gives minimal phase contrast à Donnerstag, 3. Juli 14 Positive phase contrast Phase contrast exit wave = scattered wave + unscattered wave A perfect lens in focus gives minimal phase contrast à Positive phase contrast Since ψscattered = φψunscattered with φ << 1: ψscattered << ψunscattered Iimage= |ψexit|2 =| ψunscattered + iψscattered|2 ≈ |ψunscattered|2 = Iincident Donnerstag, 3. Juli 14 Phase contrast exit wave = scattered wave + unscattered wave Ψexit = ψunscattered + iψscattered exit wave Minimal contrast Maximal contrast Defocus phase contrast Minimal contrast Lens aberrations cause an additional phase shift that generates phase contrast Donnerstag, 3. Juli 14 Lens aberrations Spherical aberration Donnerstag, 3. Juli 14 Chromatic aberration Defocus generates an additional phase shift In a perfect lens, the number of Image plane Perfect lens Donnerstag, 3. Juli 14 wavelengths from object to focal point is the same for each ray, independent of scattering angle (Fermat’s principle) Defocus generates an additional phase shift In a perfect lens, the number of Image plane Perfect lens wavelengths from object to focal point is the same for each ray, independent of scattering angle (Fermat’s principle) Lens with spherical aberration Image plane Donnerstag, 3. Juli 14 Defocus generates an additional phase shift In a perfect lens, the number of Image plane Perfect lens wavelengths from object to focal point is the same for each ray, independent of scattering angle (Fermat’s principle) Lens with spherical aberration Image plane f1 Donnerstag, 3. Juli 14 Defocus generates an additional phase shift In a perfect lens, the number of Image plane Perfect lens wavelengths from object to focal point is the same for each ray, independent of scattering angle (Fermat’s principle) Lens with spherical aberration Image plane f2 f1 Donnerstag, 3. Juli 14 Defocus generates an additional phase shift In a perfect lens, the number of Image plane Perfect lens wavelengths from object to focal point is the same for each ray, independent of scattering angle (Fermat’s principle) Lens with spherical aberration Image plane Spherical aberration causes a path length difference χ which depends on the scattering angle Donnerstag, 3. Juli 14 f2 f1 χ= e.g. 3λ/2 Depending on its phase shift, the scattered wave reinforces or weakens the exit wave exit wave = scattered wave + unscattered wave Ψexit = ψunscattered + iψscattered scattered and unscattered wave are in phase: exit wave gets stronger scattered wave is shifted by π/2 = 90° relative to unscattered wave: exit wave is nearly same as unscattered wave, therefore phase contrast is minimal scattered and unscattered wave are out of phase by π = 90°: exit wave gets weaker Donnerstag, 3. Juli 14 Optimal (Scherzer) focus The phase shift χ(θ) introduced by the spherical aberration Cs of the objective lens depends on the scattering angle θ and on the amount of defocus Δf. ! χ(θ) = (2π/λ) [Δf θ2/2 - Cs θ4/4] The spatial frequency R (measured in Å -1) is the reciprocal of the distance between any two points of the specimen. For small scattering angles, R ≈ θ / λ. With this approximation, the phase shift expressed as a function of R is χ(R) = (π/2) [2Δf R2 λ - Cs R4 λ3]! (Scherzer formula) At Scherzer focus, the terms containing Δf and Cs nearly add up to 1, and a positive phase shift of χ ≈ +π/2 applies to a wide resolution range. Donnerstag, 3. Juli 14 Defocus and spherical aberration have similar but opposite effects Donnerstag, 3. Juli 14 Contrast transfer in exact focus Scherzer focus high underfocus (Erikson and Klug, 1971) Donnerstag, 3. Juli 14 The contrast transfer function (CTF) The phase Contrast Transfer Function B(θ) of the electron microscope is defined as ! B(θ) = -2 sin χ(θ) For any particular defocus Δf, the CTF indicates the range of scattering angles θ in which the object is imaged with positive (or negative) phase contrast. For scattering angles where CTF = 0, there is no phase contrast. As spherical aberration Cs is constant, Δf can be adjusted to produce maximum phase contrast of ±π/2 for a particular range of scattering angles. Donnerstag, 3. Juli 14 Fourier transforms of amorphous carbon film images underfocus FT Donnerstag, 3. Juli 14 underfocus FT Fourier transforms of amorphous carbon film images underfocus FT underfocus FT Contrast Transfer Function, CTF Donnerstag, 3. Juli 14 CTF as a function of defocus Movie by Henning Stahlberg, UC Davis Donnerstag, 3. Juli 14 CTF as a function of defocus Movie by Henning Stahlberg, UC Davis Donnerstag, 3. Juli 14 The transmission electron microscope electron source specimen objective lens diffraction plane image plane focus plane Donnerstag, 3. Juli 14 Film image Janet Vonck, Deryck Mills Donnerstag, 3. Juli 14 DQE of direct electron detectors DQE = SNR2o/SNR2i counting mode, super-resolution McMullan, Faruqi and Henderson; http://arxiv.org/pdf/1406.1389.pdf Donnerstag, 3. Juli 14 CCD vs. direct detector 300 kV electrons 50 µm Scintillator Fiber optics CCD chip -35°C Separate vacuum conventional CCD detector Peltier cooler Falcon-II direct electron detector Richard Henderson, Greg McMullan (MRC-LMB Cambridge, UK) Donnerstag, 3. Juli 14 CCD vs. direct detector 300 kV electrons 50 µm Scintillator Fiber optics CCD chip 4000 x 4000 14 µm pixels read out 17 times per second -35°C Separate vacuum conventional CCD detector Peltier cooler Falcon-II direct electron detector Richard Henderson, Greg McMullan (MRC-LMB Cambridge, UK) Donnerstag, 3. Juli 14 Fast readout overcomes drift 3.36 Å Frame displacement in Å Matteo Allegretti, Janet Vonck, Deryck Mills Donnerstag, 3. Juli 14 Science, 28 March 2014 Donnerstag, 3. Juli 14 Science, 28 March 2014 Science, 28 March 2014 Donnerstag, 3. Juli 14 Science, 28 March 2014
